WO2002081096A1

WO2002081096A1 - Flexible centrifuge bag and methods of use

Info

Publication number: WO2002081096A1
Application number: PCT/US2002/010991
Authority: WO
Inventors: Victor D. Dolecek; David Malcolm
Original assignee: Medtronic, Inc.
Priority date: 2001-04-09
Filing date: 2002-04-05
Publication date: 2002-10-17
Also published as: EP1414581A1

Abstract

A flexible, disposable centrifuge bag (226) for receiving and holding a fluid sample, comprising one or more tubes (248, 250) and upper and lower flexible sheets (234, 236) having doughnut shaped configurations. The upper and lower sheets are superimposed and completely sealed together at their outer perimeters (238) to define an outer perimeter (240) of the centrifuge bag. The inner perimeter of the bag defines a central core (242). The tubes are sandwiched between the upper and lower sheets and extend from the central core of the centrifuge bag. The upper and lower sheets are sealed together at their inner perimeters such that the tubes are sealed between said upper and lower sheets at the inner perimeter. The bag may be used to harvest platelet rich plasma from whole blood in either a batch method or while simultaneously infusing additional aliquots of whole blood into the bag during centrifugation.

Description

FLEXIBLE CENTRIFUGE BAG AND METHODS OF USE

BACKGROUND OF THE INVENTION

Field ofthe Invention: This invention relates to novel methods, devices and apparatuses for the centrifugal separation of a liquid into its components of varying specific gravities, and is more particularly directed toward a blood separation device usefiil, for example, in the separation of blood components for use in various therapeutic regimens. Description ofthe State of Ait: Centrifugation utilizes the principle that particles suspended in solution will assume a particular radial position within the centrifuge rotor based upon their respective densities and will therefore separate when the centrifuge is rotated at an appropriate angular velocity for an appropriate period of time. Centrifugal liquid processing systems have found applications in a wide variety of fields. For example, centrifugation is widely used in blood separation techniques to separate blood into its component parts, that is, red blood cells, platelets, white blood cells, and plasma.

The liquid portion ofthe blood, referred to as plasma, is a protein-salt solution in which red and white blood cells and platelets are suspended. Plasma, which is 90 percent water, constitutes about 55 percent ofthe total blood volume. Plasma contains albumin (the chief protein constituent), fibrinogen (responsible, in part, for the clotting of blood), globulins (including antibodies) and other clotting proteins. Plasma selves a variety of functions, from maintaining a satisfactory blood pressure and providing volume to supplying critical proteins for blood clotting and immunity. Plasma is obtained by separating the liquid portion of blood from the cells suspended therein. Red blood cells (erythrocytes) are perhaps the most recognizable component of whole blood. Red blood cells contain hemoglobin, a complex iron-containing protein that canies oxygen throughout the body while giving blood its red color. The percentage of blood volume composed of red blood cells is called the "hematocrit." White blood cells (leukocytes) are responsible for protecting the body from invasion by foreign substances such as bacteria, fungi and viruses. Several types of white blood cells exist for this puipose, such as granulocytes and macrophages which protect against infection by surrounding and destroying invading bacteria and vinises, and lymphocytes which aid in the immune defense.

Platelets (thrombocytes) are veiy small cellular components of blood that help the clotting process by sticking to the lining of blood vessels. Platelets are vital to life, because they help prevent both massive blood loss resulting from trauma and blood vessel leakage that would otherwise occur in the course of normal, day-to-day activity. If whole blood is collected and prevented from clotting by the addition of an appropriate anticoagulant, it can be centrifuged into its component parts. Centrifugation will result in the red blood cells, which weigh the most, packing to the most outer portion ofthe rotating container, while plasma, being the least dense will settle in the central portion ofthe rotating container. Separating the plasma and red blood cells is a thin white or grayish layer called the buffy coat. The buffy coat layer consists ofthe white blood cells and platelets, which together make up about 1 percent ofthe total blood volume.

These blood components, discussed above, may be isolated and utilized in a wide range of diagnostic and therapeutic regimens. For example, red blood cells are routinely transfused into patients with chronic anemia resulting from disorders such as kidney failure, malignancies, or gastrointestinal bleeding and those with acute blood loss resulting from ttauma or surgery. The plasma component is typically frozen by cryoprecipitation and then slowly thawed to produce cryoprecipitated antihemophiliac factor (AHF) which is rich in certain clotting factors, including Factor VIII, fibrinogeii, von Willebrand factor and Factor XIII. Cryoprecipitated AHF is used to prevent or control bleeding in individuals with hemophilia and von Willebrand's disease. Platelets and white blood cells, which are found in the buffy layer component, can be used to treat patients with abnormal platelet function (thrombocytopenia) and patients that are unresponsive to antibiotic therapy, respectively.

Various techniques and apparatus have been developed to facilitate the collection of whole blood and the subsequent separation of therapeutic components therefrom. Centrifugal systems, also referred to as blood-processing systems, generally fall into two categories, discontinuous-flow and continuous-flow devices.

In discontinuous-flow systems, whole blood from the donor or patient flows through a conduit into the rotor or bowl where component separation takes place. These systems employ a bowl-type rotor with a relatively large (typically 200 ml or more) volume that must be filled with blood before any ofthe desired components can be harvested. When the bowl is full, the drawing of fresh blood is stopped, the whole blood is separated into its components by centrifugation, and the unwanted components are returned to the donor or patient through the same conduit intermittently, in batches, rather than on a continuous basis. When the return has been completed, whole blood is again drawn from the donor or patient, and a second cycle begins. This process continues until the required amount of the desired component has been collected.

Discontinuous-flow systems have the advantage that the rotors are relatively small in diameter but have the disadvantage that the extracorporeal volume (i.e., the amount of blood that is out ofthe donor at any given time during the process) is large. This, in turn, makes it difficult or impossible to use discontinuous systems on people whose size and weight will not pennit the drawing ofthe amount of blood required to fill the rotor. Discontinuous-flow devices are used for the collection of platelets and/or plasma, and for the concenfration and washing of red blood cells. They are used to reconstitute previously frozen red blood cells and to salvage red blood cells lost intraoperatively. Because the bowls in these systems are rigid and have a fixed volume, however, it is difficult to confrol the hematocrit ofthe final product, particularly ifthe amount of blood salvaged is insufficient to fill the bowl with red blood cells.

One example of a discontinuous-flow system is disclosed by McMannis, et al, in his U.S. Patent No. 5,316,540, and is a variable volume centrifuge for separating components of a fluid medium, comprising a centrifuge that is divided into upper and lower chambers by a flexible membrane, and a flexible processing container bag positioned in the upper chamber of the centrifuge. The McMannis, et al, system varies the volume ofthe upper chamber by pumping a hydraulic fluid into the lower chamber, which in turn raises the membrane and squeezes the desired component out ofthe centrifuge. The McMannis, et al, system takes up a fairly large amount of space, and its flexible pancake-shaped rotor is awkward to handle. The McMannis, et al, system does not permit the fluid medium to flow into and out of the processing bag at the same time, nor does it permit fluid medium to be pulled out ofthe processing bag by suction.

In continuous-flow systems, whole blood from the donor or patient also flows through one conduit into the spinning rotor where the components are separated. The component of interest is collected and the unwanted components are returned to the donor through a second conduit on a continuous basis as more whole blood is being drawn. Because the rate of drawing and the rate of return are substantially the same, the extracorporeal volume, or the amount of blood that is out ofthe donor or patient at any given time in the procedure, is relatively small. These systems typically employ a belt- type rotor, which has a relatively large diameter but a relatively small (typically 100 ml or less) processing volume. Although continuous-flow systems have the advantage that the amount of blood that must be outside the donor or patient can be relatively small, they have the disadvantage that the diameter of the rotor is large. These systems are, as a consequence, large. Furthermore, they are complicated to set up and use. These devices are used almost exclusively for the collection of platelets.

Continuous-flow systems are comprised of rotatable and stationary parts that are in fluid communication. Consequently, continuous-flow systems utilize either rotary seals or a J-loop. A variety of types of rotaiy centrifuge seals have been developed. Some examples of rotary centrifuge seals which have proven to be successful are described in

U.S. Pat. Nos. 3,409,203 and 3,565,330, issued to Latham. In these patents, rotary seals are disclosed which are fonned from a stationary rigid low friction member in contact with a moving rigid member to create a dynamic seal, and an elastomeric member which provides a resilient static seal as well as a modest closing force between the surfaces ofthe dynamic seal.

Another rotary seal suitable for use in blood-processing centrifuges is described in U.S. Pat. No. 3,801,142 issued to Jones, et al. In this rotary seal, a pair of seal elements having confronting annular fluid-tight sealing surfaces of non-con-odible material are provided. These are maintained in a rotatable but fluid-tight relationship by axial compression of a length of elastic tubing forming one of the fluid connections to these seal elements. Related types of systems which incorporate rotatable, disposable annular separation chambers coupled via rotary seals to stationary tubing members are disclosed in U.S. Pat. Nos. 4,387,848; 4,094,461; 4,007,871; and 4,010,894.

One drawback present in the above-described continuous-flow systems has been their use of a rotating seal or coupling element between that portion ofthe system carried by the centrifuge rotor and that portion ofthe system which remains stationary. While such rotating seals have provided generally satisfactory performance, they have been expensive to manufacture and have unnecessarily added to the cost ofthe flow systems. Furthermore, such rotating seals introduce an additional component into the system which if defective can cause contamination of the blood being processed.

One flow system heretofore contemplated to overcome the problem ofthe rotating seal utilizes a rotating carriage on which a single housing is rotatably mounted. An umbilical cable extending to the housing from a stationary point imparts planetary motion to the housing and thus prevents the cable from twisting. To promote the desired ends of sterile processing and avoid the disadvantages of a discontinuous-flow system within a single sealed system, a family of dual member centrifuges can be used to effect cell separation. One example of this type of centrifuge is disclosed in U.S. Pat. No. RE 29,738 to Adams entitled "Apparatus for Providing Energy Communication Between a Moving and a Stationary Terminal". As is now well known, due to the characteristics of such dual member centrifuges, it is possible to rotate a container containing a fluid, such as a unit of donated blood and to withdraw a separated fluid component, such as plasma, into a stationary container, outside of the centrifuge without using rotating seals. Such container systems utilize a J-loop and can be formed as closed, sterile transfer sets.

The Adams patent discloses a centrifuge having an outer rotatable member and an inner rotatable member. The inner member is positioned within and rotatably supported by the outer member. The outer member rotates at one rotational velocity, usually called "one omega," and the im er rotatable member rotates at twice the rotational velocity ofthe outer housing or "two omega." There is thus a one omega difference in rotational speed of the two members. For puiposes of this document, the tenn "dual member centrifuge" shall refer to centrifuges ofthe Adams type. The dual member centrifuge ofthe Adams patent is particularly advantageous in that, as noted above, no seals are needed between the container of fluid being rotated and the non-moving component collection containers. The system ofthe Adams patent provides a way to process blood into components in a single, sealed, sterile system wherein whole blood from a donor can be infused into the centrifuge while the two members ofthe centrifuge are being rotated.

An alternate to the apparatus ofthe Adams patent is illusfrated in U.S. Pat. No. 4,056,224 to Lolachi entitled "Flow System for Centrifugal Liquid Processing Apparatus." The system ofthe Lolachi patent includes a dual member centrifuge ofthe Adams type. The outer member ofthe Lolachi centrifuge is rotated by a single electric motor which is coupled to the internal rotatable housing by belts and shafts.

U.S. Pat. No. 4,108,353 to Brown entitled "Centrifugal Apparatus With Oppositely Positioned Rotational Support Means" discloses a centrifuge structure ofthe Adams type which includes two separate electrical motors. One electric motor is coupled by a belt to the outer member and rotates the outer member at a desired nominal rotational velocity.

The second motor is carried within the rotating exterior member and rotates the inner member at the desired higher velocity, twice that ofthe exterior member.

U.S. Pat. No. 4,109,855 to Brown, et al, entitled "Drive System For Centrifugal Processing Apparatus" discloses yet another drive system. The system ofthe Brown, et al. , patent has an outer shaft, affixed to the outer member for rotating the outer member at a selected velocity. An inner shaft, coaxial with the outer shaft, is coupled to the inner member. The inner shaft rotates the inner member at twice the rotational velocity as the outer member. A similar system is disclosed in U.S. Pat. No. 4,109,854 to Brown entitled "Centrifugal Apparatus With Outer Enclosure". The continuous-flow systems described above are large and expensive units that are not intended to be portable. Further, they are also an order of magnitude more expensive than a standard, multi-container blood collection set. There exists the need, therefore, for a centrifugal system for processing blood and other biological fluids that is compact and easy to use and that does not have the disadvantages of prior-art continuous- flow systems.

Whole blood that is to be separated into its components is commonly collected into a flexible plastic donor bag, and the blood is centrifuged to separate it into its components through a batch process. This is done by spinning the blood bag for a period of about 10 minutes in a large refrigerated centrifuge. The main blood constituents, i.e., red blood cells, platelets and white cells, and plasma, having sedimented and formed distinct layers, are then expressed sequentially by a manual extractor in multiple satellite bags attached to the primary bag.

More recently, automated extractors have been introduced in order to facilitate the manipulation. Nevertheless, the whole process remains laborious and requires the separation to occur within a certain time frame to guarantee the quality of the blood components. This complicates the logistics, especially considering that most blood donations are performed in decentralized locations where no batch processing capabilities exist.

This method has been practiced since the widespread use ofthe disposable plastic bags for collecting blood in the 1970's and has not evolved significantly since then. Some attempts have been made to apply haemapheresis technology in whole blood donation.

This technique consists of drawing and extracting on-line one or more blood components while a donation is performed, and returning the remaining constituents to the donor. However, the complexity and costs of haemapheresis systems preclude their use by transfusion centers for routine whole blood collection. There have been various proposals for portable, disposable, centrifugal apparatus, usually with collapsible bags, for example as in U.S. Pat. Nos. 3,737,096, or 4,303,193 to Latham, Jr., or with a rigid walled bowl as in U.S. Pat. No. 4,889,524 to Fell, et al. These devices all have a minimum fixed holding volume which requires a minimum volume usually of about 250 ml to be processed before any components can be collected. U.S. Pat. No. 5,316,540 to McMannis, et al, discloses a centrifugal processing apparatus, wherein the processing chamber is a flexible processing bag which can be deformed to fill it with biological fluid or empty it by means of a membrane which forms part ofthe drive unit. The bag comprises a single inlet/outlet tubing for the introduction and removal of fluids to the bag, and consequently cannot be used in a continual, on-line process. Moreover, the processing bag has a the disadvantage of having 650 milliliter capacity, which makes the McMannis, et al, device difficult to use as a blood processing device.

As discussed above, centrifuges are often used to separated blood into its components for use in a variety of therapeutic regimens. One such application is the preparation of a bioadhesive sealant. A bioadhesive sealant, also refened to as a fibrin glue, is a relatively new technological advance which attempts to duplicate the biological process ofthe final stage of blood coagulation. Clinical reports document the utility of fibrin glue in a variety of surgical fields, such as, cardiovascular, thoracic, transplantation, head and neck, oral, gastrointestinal, orthopedic, neurosurgical, and plastic surgeiy. At the time of surgery, the two primary components comprising the fibrin glue, fibrinogen and thrombin, are mixed together to form a clot. The clot is applied to the appropriate site, where it adheres to the necessaiy tissues, bone, or nerve within seconds, but is then slowly reabsorbed by the body in approximately 10 days by fibrinolysis. Important features of fibrin glue is its ability to: (1) achieve haemostasis at vascular anastomoses particularly in areas which are difficult to approach with sutures or where suture placement presents excessive risk; (2) control bleeding from needle holes or arterial tears which cannot be controlled by suturing alone; and (3) obtain haemostasis in heparinized patients or those with coagulopathy. See, Borst, H.G., et al, JThorac. Cardiovasc. Surg., 84:548-553 (l^c\2); Walterbusch, G. J et al., Tliorac. Cardiovasc. Surg., 30:234-23 5 (1982); and Wolner, F. J, et al, Thorac. Cardiovasc. Surg., 30:236-237 (1982). Despite the effectiveness and successful use of fibrin glue by medical practitioners in Europe, neither fibrin glue nor its essential components fibrinogen and thrombin are widely used in the United States. In large part, this stems from the 1978 U.S. Food and Drag Administration ban on the sale of commercially prepared fibrinogen concentrate made from pooled donors because ofthe risk of transmission of viral infection, in particular the hepatitis-causing viruses such as HBV and HCV (also known as non-A and non-B hepatitis virus). In addition, the more recent appearance of other lipid-enveloped viruses such as HIV, associated with AIDS, cytomegalovirus (CMV), as well as Epstein-Baιτ virus and the herpes simplex viruses in fibrinogen preparations makes it unlikely that there will be a change in this policy in the foreseeable future. For similar reasons, human thrombin is also not curcenfly authorized for human use in the United

States. Bovine thrombin, which is licensed for human use in the United States is obtained from bovine sources which do not appear to carry significant risks for HIV and hepatitis, although other bovine pathogens, such as bovine spongifonn and encephalitis, may be present.

There have been a variety of methods developed for preparing fibrin glue. For example, Rose, et al. in U.S. Pat. No. 4,627,879 discloses a method of preparing a ciyoprecipitated suspension containing fibrinogen and Factor XIII useful as a precursor in the preparation of a fibrin glue which involves (a) freezing fresh frozen plasma from a single donor such as a human or other animal, e.g. a cow, sheep or pig, which has been screened for blood transmitted diseases, e.g. one or more of syphilis, hepatitis or acquired immune deficiency syndrome, at about 80° C for at least about 6 hours, preferably for at least about 12 hours; (b) raising the temperature ofthe frozen plasma, e.g. to between about 0° C and room temperature, so as to form a supernatant and a cryoprecipitated suspension containing fibrinogen and Factor XIII; and (c) recovering the cryoprecipitated suspension. The fibrin glue is then prepared by applying a defined volume ofthe cyroprecipitate suspension described above and applying a composition containing a sufficient amount of tlirombin, e.g. human, bovine, ovine or porcine tlirombin, to the site so as to cause the fibrinogen in the suspension to be converted to the fibrin glue which then solidifies in the form of a gel.

A second technique for preparing fibrin glue is disclosed by Marx in his U.S. Patent Number 5,607,694. Essentially, a cryoprecipitate as discussed previously selves as the source of the fibrinogen component and then Marx adds thrombin and liposomes. A third method discussed by Bemiyer, (M.,) et al, entitled "Immunization by bovine thrombin used with fibrin glue during cardiovascular operations," (J.) T orac. Cardiovasc. Surg., 105(5):892-897 (1992)) discloses a fibrin glue prepared by mixing bovine tlirombin not only with human coagulant proteins, such as fibrinogen, fibronectin, Factor XIII, and plasminogen, but also with bovine aprotinin and calcium chloride.

The above patents by Rose, et al, and Marx, and the technical paper by Benuyer, et al. each disclose methods for preparing fibrin sealants; however, each of these methods suffer disadvantages associated with the use of bovine tlirombin as the activating agent. A serious and life threatening consequence associated with the use of fibrin glues comprising bovine thrombin is that patients have been reported to have a bleeding diathesis after receiving topical bovine thrombin. This complication occurs when patients develop antibodies to the bovine factor V in the relatively impure bovine tlirombin preparations. These antibodies cross-react with human factor V, thereby causing a factor V deficiency that can be sufficiently severe to induce bleeding and even death. See, Rapaport, S.I., et al, Am.(J.) Clin. Pathol, 97:84-91 (1992); Berruyer, M., et al, J. Thorac. Cardiovasc.

Surg., 105:892-897 (1993); Zehnder, J., et l, Blood, 76(10):2011-2016 (1990); Muntean, W., et al, Acta Paediatr., 83:84-7 (1994); Christine, R.J., et al, Surgeiy, 127:708-710 (1997).

Further disadvantages associated with the methods disclosed by Marx and Rose, et al. are that the ciyoprecipitate preparations require a large time and monetary commitment to prepare. Furthermore, great care must be taken to assure the absence of any viral contaminants.

A further disadvantage associated with the methods previously disclosed is that while human thrombin is contemplated for use as an activator, human thrombin is not available for clinical use and there is no evidence that patients will not have an antigenic response to human thrombin. By analogy, recombinant human factor VHI has been shown to produce antigenic responses in hemophiliacs. See, Biasi, R. de., Thrombosis and Haemostasis, 71(5):544-547 (1994). Consequently, until more clinical studies are performed on the effect of human recombinant thrombin one cannot merely assume that the use of recombinant human thrombin would obviate the antigenic problems associated with bovine thrombin. A second difficulty with thrombin is that it is autocatalytic, that is, it tends to self-destruct, making handling and prolonged storage a problem.

Finally, as discussed above, fibrin glue is comprised primarily of fibrinogen and thrombin thus lacking an appreciable quantity of platelets. Platelets contain growth factors and healing factors which are assumed to be more prevalent in a platelet concentrate.

Moreover, platelets aid in acceleration ofthe clotting process.

There is still a need, therefore, for a centrifugal system for processing blood and other biological fluids, that is compact and easy to use and that does not have the disadvantages of prior-art continuous-flow systems and furthermore there exists a need for a convenient and practical method for preparing a platelet gel composition wherein the resulting platelet gel poses a zero risk of disease transmission and a zero risk of causing an adverse physiological reaction.

There is also a widespread need for a system that, during blood collection, will automatically separate the different components of whole blood that are differentiable in density and size, with a simple, low cost, disposable unit.

There is further a need for a centrifugal cell processing system wherein multiple batches of cells can be simultaneously and efficiently processed without the use of rotational coupling elements.

There is yet a further need for a platelet concentrate that aids in increasing the rate of fibrin clot formation, thereby facilitating haemostasis.

Preferably the apparatus will be essentially self-contained. Preferably, the equipment needed to practice the method will be relatively inexpensive and the blood contacting set will be disposable each time the whole blood has been separated.

SUMMARY OF THE INVENTION Accordingly, one object of this invention is to provide a method and apparatus for the separation of components suspended or dissolved in a fluid medium by cenhifugation. More specifically, one object of this invention is to provide a method for the separation and isolation of one or more whole blood components, such as platelet rich plasma, white blood cells and platelet poor plasma, from anticoagulated whole blood by cenhifugation, wherein the components are isolated while the centrifuge is rotating.

Another object of this invention is to utilize the isolated cell components in a therapeutic regimen.

Another object of this invention is to provide an apparatus for the separation of whole blood components, wherein the apparatus contains a centrifuge bag that provides for simultaneous addition of whole blood from a source container and the withdrawal of a specific blood component during centrifugation.

Another object of this invention is to provide disposable, single-use centrifuge bags for holding whole blood during the separation of components ofthe whole blood by centrifugation, wherein the bag is adapted for use in a portable, point-of-use centrifuge. Another object of this invention is to provide a portable centrifuge containing a disposable centrifuge bag that maximizes d e amount of a predetermined blood fraction that can be harvested from an aliquot of blood that is of greater volume than the capacity ofthe disposable centrifuge bag.

To achieve the foregoing and other objects and in accordance with the puiposes of the present invention, as embodied and broadly described therein, one embodiment of this invention comprises a flexible, disposable centrifuge bag adapted to be rotated about an axis, comprising: a) one or more tubes, and b) upper and lower flexible sheets, each sheet having a doughnut shaped configuration, an inner perimeter defining a central core and an outer perimeter, wherein the upper and lower sheets are superimposed and completely sealed together at their outer perimeters, and wherein the tubes are sandwiched between the upper and lower sheets and extend from the central core toward the outer perimeter, such that when the upper and lower sheets are sealed at the inner perimeter the tubes are sealed between the upper and lower sheets at the inner perimeter and are in fluid communication with the environment inside and outside the centrifuge bag. The one or more tubes are fluidly connected to an umbilical cable comprising one or more lumen equal to the number of tabes ofthe centrifuge bag.

To further achieve the foregoing and other objects of this invention, another embodiment ofthe present invention comprises a rigid molded container adapted to be rotated about an axis, comprising a rigid, annular body having an axial core that is closed at the top end and opened at the bottom end. The rigid molded container further comprises an interior collection chamber for receiving and holding a fluid medium to be centrifuged, the chamber having an outer perimeter, an inner perimeter, and a generally off-centered "figure eight" shaped cross-sectional area. The rigid molded container further comprises a first channel which extends radially from the core and is in fluid communication with a point near the outer perimeter ofthe chamber, and a second channel which extends radially from the core and is in fluid communication with an area near the nairow portion or "neck" ofthe figure eight-shaped chamber. The first and second channels thus provide fluid communication with the environment inside and outside the interior collection chamber. The first and second channels are fluidly connected to a dual lumen tubing having an inlet lumen and an outlet lumen. To further achieve the foregoing and other objects of this invention, another embodiment ofthe present invention is an apparatus and method for separating components contained in a fluid medium. More particularly, the present invention utilizes the principles of centrifugation to alloλv for the separation of whole blood into fractions such as platelet rich plasma and platelet poor plasma, hi one aspect ofthe present invention, the above-described separation ofthe components is provided by utilizing a rotatable centrifuge motor comprismg a base having a cenfral column and a disposable centrifuge bag having a central core and which is positionable within the centriftige motor and rotatable therewith. The disposable centrifuge bag, which holds the whole blood during centrifugation, further comprises an inlet tube for introducing the whole blood to the centrifuge bag, and an outlet tube for removing the desired blood fraction from the centrifuge bag. The inlet and outlet tubes are in fluid communication with a dual lumen tubing. The centrifuge bag is removably fixed within the centrifuge rotor by inserting the raised column through the bag center core and securing with the cover. During the rotation ofthe centrifuge, components ofthe whole blood will assume a radial, horizontal position within the centrifuge bag based upon a density of such components, and thus the fluid medium components will be separated from other components having different densities.

Once a desired degree of separation of whole blood has been achieved, the present invention provides for the specific removal ofthe desired fraction within one or more of the regions from the centrifuge bag through the outlet tube during continued rotation o the centriftige, thereby allowing for on-line removal ofthe desired fraction. Additional aliquots may be added to the centrifuge bag via the inlet tube simultaneously or after the desired component has been harvested. In one embodiment, the centrifuge bag is a flexible, transparent, generally flat doughnut-shaped bag. In another embodiment, the centrifuge bag is a rigid, transparent container having an interior chamber for receiving and holding the fluid medium during centrifugation, the interior chamber having a generally off-centered figure eight cross-sectional configuration.

Another aspect ofthe present invention comprises a disposable centrifuge bag having an inlet tube and an outlet tube, wherein the outlet tube is fluidly connected with a bent fitting. Another aspect ofthe present invention comprises a centrifuge rotor for holding a centrifuge bag, the rotor comprising a base and a cover, the base further having a first grooved, raised center column and the cover having a second grooved, raised center column. The centrifuge bag is a flexible, douglmut-shaped bag comprising inlet and outlet tubes in fluid communication with the environment inside and outside the centrifuge bag, wherein the tabes are seated in the base and cover column grooves to hold the centriftige bag in a fixed position relative to the base and cover, such that the bag does not spin independently ofthe base and cover but rather spins concurrently and at the same rate of rotation as the base and cover. Another aspect ofthe present invention comprises a centriftige rotor for holding a centrifuge bag, the rotor comprising a base and a cover for securing a centrifuge bag therebetween, the centriftige cover further comprising one or more concentric indicator circles that are spaced from the center of the cover or the base to aid the operator in visualizing the distal ends of these tabes. Another aspect ofthe present invention for the separation of components of a fluid medium (e.g., whole blood) utilizes a centrifuge rotor comprising an interior chamber having a complex configuration, wherein the chamber holds a flexible, doughnut-shaped centrifuge bag for retaining the fluid medium during centrifugation. The centrifuge rotor is defined by a base having a lower chamber, and a cover having an upper chamber. When the cover is superimposed on the base, the upper and lower chambers define the annular interior chamber of the rotor. The interior rotor chamber has a generally off-centered figure eight-shaped cross-sectional configuration specifically designed to maximize the collection of the desired component (e.g., platelet rich plasma) by centrifiigation of a fluid medium (e.g., anticoagulated whole blood). The centrifuge bag is formed from a substantially flexible material, such that the profile of the centrifuge bag during cenhifugation is thus detennined at least in part by the volume ofthe fluid medium contained therein. When the centrifuge bag is filled to maximum capacity, it assumes the configuration of the interior of the rotor chamber.

Another aspect of this invention comprises a method for on-line harvesting of a predetermined component of a fluid medium. One embodiment of the present invention utilizes a centrifuge and a disposable centrifuge bag for containing the fluid medium during separation and which is positionable within the centrifuge, the centrifuge bag further comprising at least one inlet tube and at least one outlet tube. The centrifuge includes a centrifuge rotor having a base portion, a cover, and an outer rim. The base portion and the cover define the interior ofthe centrifuge rotor, which is separated into upper and lower chambers. The disposable centrifuge bag is positionable horizontally within the lower chamber and may be appropriately secured to the centrifuge base by the cover. The centriftige bag is fluidly connected via a dual lumen tubing to a source (e.g., to a container comprising anticoagulated autologous whole blood) and collection container (e.g., for receiving platelet rich plasma or some other component that will then be ftirther processed). The dual lumen tubing comprises an inlet lumen fluidly connected to the inlet tube o the centrifuge bag and an outlet lumen fluidly connected to the outlet tube ofthe centrifuge bag. The cenfrifuge bag is substantially annular relative to the rotational axis of the centrifuge. When the centrifuge bag is positioned within the centrifuge rotor and appropriately secured thereto to allow for simultaneous rotation, the fluid medium may be provided to the centrifuge bag via the inlet lumen of the tubing during rotation ofthe centrifuge. The components ofthe bag assume radial, horizontal positions base based on their densities. When a desired degree of separation has been achieved, the desired fraction may be removed from the centrifuge bag via the outlet lumen during continued rotation ofthe cenfrifuge. The position ofthe fraction to be harvested may be shifted into the area of the outlet tabe as needed, either by withdrawing components that are positioned near the outer perimeter through the inlet tabe, or by adding additional aliquots of the fluid medium to the bag. In one embodiment of this method, the bag is a flexible, transparent doughnut-shaped bag. In another embodiment of this method, the bag is a rigid, transparent bag comprising an interior chamber having an off-centered, figure eight cross- sectional configuration.

It is yet another object ofthe invention to provide a centrifugal liquid processing system that may be automated.

It is yet another object ofthe present invention to provide a centrifuge having an internal lead drive mechanism allowing for a compact size. A further object of the present invention is to provide for a method and device for the production and isolation of thrombin for all medical uses. It is yet another object of this invention to provide a method for preparing a completely autologous platelet gel.

Another object ofthe present invention is to provide an autologous platelet gel wherein the risks associated with the use of bovine and recombinant human thrombin are eliminated.

A further object ofthe present invention is to provide an autologous platelet gel for any application.

It is a further object ofthe present invention to provide cellular components to be used in medical applications. Additional objects, advantages, and novel features of this invention shall be set forth in part in the description and examples that follow, and in part will become apparent to those skilled in the art upon examination ofthe following or may be learned by the practice ofthe invention. The objects and the advantages ofthe invention may be realized and attained by means ofthe instrumentalities and in combinations paiticularly pointed out in the appended claims.

BRIEF DESCRIPTION OF THE DRAWINGS

The accompanying drawings, which are incorporated in and form a part ofthe specifications, illusfrate the preferred embodiments ofthe present invention, and together with the description serve to explain the principles ofthe invention. In the Drawings:

Figure 1 is a perspective view illusfrating one embodiment ofthe continuous-flow centrifugal processing system ofthe present invention illusfrating a centrifuge and side- mounted motor positioned within a protective housing or enclosure ofthe invention.

Figure 2 is an exploded side view ofthe centrifuge and the side-mounted motor of the centrifugal processmg system of Figure 1 illustrating the individual components ofthe centrifuge.

Figure 3 is a partial perspective view ofthe lower case assembly ofthe drive shaft assembly of Figure 2.

Figure 4 is an exploded side view ofthe lower case assembly of Figure 3. Figure 5 is an exploded perspective view ofthe components ofthe lower case assembly of Figure 3.

Figure 6 is a top view ofthe lower bearing assembly which is positioned within the lower case assembly of Figure 3.

Figure 7 is a perspective view ofthe lower bearing assembly of Figure 6. Figure 8 is an exploded side view ofthe lower bearing assembly of Figures 6 and

7.

Figure 9 is a perspective view ofthe receiving tube guide ofthe centrifuge of Figure 2.

Figure 10 is an exploded, perspective view of a gear ofthe mid-shaft gear assembly of Figure 2.

Figure 11 is a perspective view ofthe gear of Figure 10 as it appears assembled. Figure 12 is an exploded, perspective view of the top bearing assembly ofthe centriftige of Figure 2.

Figure 13 is a perspective view of the top case shell ofthe top bearing assembly of Figure 12. Figure 14 is a perspective view ofthe centrifuge ofthe present invention shown in Figure 1, having a quarter section cut away along lines 14-14 of Figure 1.

Figure 15 is a perspective view of one embodiment of a centrifuge rotor base.

Figure 16 is a perspective view of one embodiment of a centrifuge rotor cover. Figure 17 is a side cross-sectional view of one embodiment of a rotor of this invention taken along view lines 17 of Figure 14 for holding a disposable centriftige bag, showing a dual lumen tubing connected to the bag.

Figure 18 is a side cross-sectional view of one embodiment of a rotor of this invention taken along view lines 18 of Figure 1 for holding a disposable centiifiige bag, showing the grooved columns ofthe base and cover.

Figure 19 is an enlarged perspective view similar to Figure 1 illusfrating an alternate embodiment of a centrifuge driven by a side-mounted motor (with only the external drive belt shown).

Figure 20 is a cutaway side view ofthe centiifiige of Figure 19 illustrating the internal pulley drive system utilized to achieve a desired drive ratio and illusfrating the rotor base configured for receiving a centrifuge bag.

Figure 21 is a cutaway side view similar to Figure 20 with the rotor base removed to better illustrate the top pulley and the location of both idler pulleys relative to the installed internal drive belt. Figure 22 is a sectional view ofthe centriftige of Figure 20 further illustrating the internal pulley drive system an showing the routing ofthe centrifuge tube (or umbilical cable).

Figure 23 is a top view of a further alternate centrifuge similar to the cenfrifuge of Figure 19 but including internal, separate bearing members (illusfrated as four cam followers) that allows the inclusion of guide shaft to be cut through portions of the cenfrifuge for positioning ofthe cenfrifuge tabe (or umbilical cable).

Figure 24 is a perspective view similar to Figure 19 illusfrating the centrifuge embodiment of Figure 23 further illusfrating the guide slot and showmg that the centiifiige can be driven by an external drive belt. Figure 25 is a top view of a flexible, disposable centrifuge bag of this invention. Figure 26 is a perspective view of a flexible, disposable centrifuge bag of this invention.

Figures 27, 28, 29, and 30 are illustrations of bent fittings of this invention having "T" shaped, "curved T" shaped, "L" shaped, and "J" shaped configurations, respectively. Figure 31 is an illustration of an inlet and/or outlet tube of this invention.

Figure 32 is a top view of a disposable cenfrifuge bag of this invention after the centrifugation of whole blood, showing the separated blood components.

Figures 33-39 are schematic illustrations of one method of this invention for separating whole blood components using a disposable centrifuge bag of this invention. Figure 40 is a top view of an alternate embodiment of a disposable cenfrifuge bag of the present invention having inner and outer chambers.

Figure 41 is a top view of the disposable centriftige bag shown in Figure 34 illusfrating movement ofthe red blood cell layer from the outer perimeter toward the inner perimeter. Figure 42 is a bottom view of an alternate embodiment of a disposable centrifuge bag of the present invention having inner and outer chambers in fluid communication with outlet and inlet ports.

Figure 43 is a side cross-sectional view of a rigid disposable cenfrifuge bag of this invention. Figure 44 is a schematic illustration of separated blood components contained in a cenfrifuge bag having an elliptical cross-sectional view of the cenfrifuge bag shown in Figure 43.

Figure 45 is a side cross-sectional view of a rigid disposable centrifuge bag of this invention. Figure 46 is a schematic illustration of the surface areas and various dimensions of the figure eight configuration as shown in Figure 45.

Figure 47 is a schematic illustration of separated blood components contained in a centiifiige bag having a figure eight side cross-sectional configuration.

Figure 48 is a side cross-sectional view of an alternative embodiment of an assembled centrifuge rotor of this invention comprising the rotor cover of Figure 49 and the rotor base of Figure 50. Figure 49 is a side cross-sectional view of an alternative embodiment of a rotor cover of this invention.

Figure 50 is a side cross-sectional view of an alternative embodiment of a rotor base of this invention. Figure 51 is a perspective view of the rotor base of Figure 50.

Figure 52 is a perspective view ofthe rotor cover of Figure 49.

Figure 53 is a block diagram illusfrating the components of a centrifugal processing system ofthe present invention.

Figure 54 is a graph illusfrating the timing and relationship of transmission of confrol signals and receipt of feedback signals during operation of one embodiment ofthe automated centrifugal processing system of Figure 53.

Figure 55 is a side view of an alternative embodiment ofthe automated centrifugal processing system of Figure 53 showing a centrifuge having a rotor wherein the reservoir extends over the outer diameter ofthe centrifuge portion that facilitates use of an externally-positioned sensor assembly.

Figure 56 is a side view of a further alternative embodiment ofthe external sensor assembly feature ofthe centrifugal processing system ofthe invention without an extended rotor and illusfrating the positioning of a reflector within the centrifuge.

Figure 57 is a side view of yet another embodiment ofthe external sensor assembly feature ofthe centrifugal processing system ofthe invention illustrating a single radiant energy source and detector device.

Figure 58 is a block diagram of a an automated centrifugal processing system, similar to the embodiment of Figure 47, including components forming a temperature control system for controlling temperatures of separated and processed products. Figure 59 is a perspective view of components ofthe temperature control system of

Figure 58.

Figure 60 is schematic and sectional view ofthe dispenser ofthe present invention.

Figure 61 is a flow diagram representing the method for isolating platelet rich plasma and platelet poor plasma for use in preparing a platelet gel ofthe present invention. Figure 62 is a flow diagram representing the final portion ofthe method for preparing a platelet gel ofthe present invention using platelet rich plasma as a starting material.

Figure 63 is a flow diagram representing the final portion ofthe method for preparing a platelet gel of the present invention using platelet poor plasma as a starting material.

Figure 64 is a graphic representation ofthe effect that the serum-to-plasma ratio has on clotting times.

Figure 65 graphically represents the effect of calcium addition on the clotting times of platelet rich plasma and platelet poor plasma.

Figure 66 is a graphic representation ofthe relationship between clotting time and actual gel time using blood drawn from a donor.

Figure 67 is a graphic representation ofthe relationship between clotting time and actual gel time using blood drawn from a donor. Figure 68 graphically represents the effect of calcium addition on clotting times and gel times using blood drawn from a donor.

Figure 69 graphically represents the effect of calcium addition on clotting times and gel times using blood drawn from a donor.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT The centrifugal processing system 10 of the present invention is best shown in

Figure 1 having a stationaiy base 12, a centrifuge 20 rotatably mounted to the stationaiy base 12 for rotation about a predetermined axis A, a rotor 202 for receiving a disposable bag (not shown) designed for continuous-flow. As illustrated, the centriftigal processing system 10 includes a protective enclosure 11 comprising the main table plate or stationaiy base 12, side walls 13, and a removable lid 15 made of clear or opaque plastic or other suitable materials to provide structural support for components ofthe centriftigal processing system 10, to provide safety by enclosing moving parts, and to provide a portable centriftigal processing system 10. The centrifugal processing system 10 further includes a clamp 22 mounted over an opening (not shown) in the lid 15. Clamp 22 secures at a point at or proximately to axis A without pinching off the flow of fluid that travels through umbilical cable 228. A side mounted motor 24 is provided and connected to the cenfrifuge 20 by way of a drive belt 26 for rotating the drive shaft assembly 28 (see Figure 2) and the interconnected and driven rotor assembly 200 in the same rotational direction with a speed ratio selected to control binding of umbilical cable 228 during operation of the system, such as a speed ratio of 2:1 (i.e., the rotor assembly 200 rotates twice for each rotation ofthe drive shaft assembly 28). The present invention is further directed toward a dispensing device 902, best shown in Figure 60 for the withdrawal and manipulation of specific blood components for various therapeutic regimens, such as but not limited to the production of platelet rich plasma, platelet poor plasma, and white blood cells which may be used for the production of autologous thrombin and autologous platelet gels. Referring now to Figure 2, the continuous-flow centrifugal processing system 10 comprises a centriftige 20 to which a rotor 202 is removably or non-removably attached. The design of centrifuge 20 and its self-contained mid-shaft gear assembly 108 (comprised of gears 110, 110', 131, and 74) is a key component ofthe invention thereby allowing for the compact size ofthe entire centrifugal processing system 10 and providing for a desired speed ratio between the drive shaft assembly 28 and the rotor assembly 200.

The centrifuge 20 is assembled, as best seen in Figure 2, by inserting the lower bearing assembly 66 into lower case shell 32 thus resulting in lower case assembly 30. Cable guide 102 and gears 110 and 110' are then positioned within lower case assembly 30, as will be discussed in more detail below, so that gears 110 and 110' are moveably of engaged with lower bearing assembly 66. Upper bearing assembly 130 is then inserted within top case shell 126 thus resulting in bearing assembly 124 which is then mated to lower case assembly 30, such that gears 110 and 110' are also moveably engaged with upper bearing assembly 130, and held in place by fasteners 29. Lower bearing assembly 66 is journaled to stationary base or main table plate 12 by screws 14, thus allowing centrifuge 20 to rotate along an axis A, perpendicular to main table plate 12 (as shown in

Figure 1).

Referring now to Figures 3, 4, and 5, the lower case assembly 30 is preferably, but not necessarily, machined or molded from a metal material and includes a lower case shell 32, timing belt ring 46, timing belt flange 50, and bearing 62 (e.g., ball bearings and the like). Lower case shell 32 includes an elongated main body 40 with a smaller diameter neck portion 36 extending from one end ofthe main body 40 for receiving timing belt ring 46 and timing belt flange 50. The larger diameter main body 40 terminates into the neck portion 36 thereby forming an external shoulder 38 having a bearing surface 42 for timing belt ring 46. Timing belt ring 46 and timing belt flange 50, as best seen in Figure 5, have inner diameters that are slightly larger than the outer diameter of neck portion 36 allowing both to fit over neck portion 36. Shoulder 38 further contains at least one and preferably four internally thread holes 44 that align with hole guides 48 and 52 in timing belt ring 46 and timing belt flange 50, respectively (shown in Figure 5). Consequently, when assembled, screws 54 are received by hole guides 52 and 48 and are threaded into thread holes 44 thus securing timing belt 46 and timing belt flange 50 onto neck portion 36. Lower case shell 32 also has an axial or sleeve bore 56 extending there through, and an internal shoulder 58, the upper surface 60 of which is in approximately the same horizontal plane as external shoulder 38. Bearing 62 (shown in Figure 4) is press fit concentrically into sleeve bore 56 so that it sits flush with upper surface 60. Internal shoulder 58 also has a lower weight bearing surface 64 which seats on the upper surface 68 of lower bearing assembly 66, shown in Figures 6-8.

Lower bearing assembly 66 comprises a lower gear insert 70, ball bearings 84, gear 74 and spring pins 76 and 76'. As will become clear, the gear 74 may be of any suitable gear design for transfening an input rotation rate to a mating or contacting gear, such as the gears 110, 110' ofthe mid-shaft gear assembly 108, with a size and tooth number selected to provide a desired gear train or speed ratio when combined with contacting gears. For example, the gear 74 may be configured as a straight or spiral bevel gear, a helical gear, a worm gear, a hypoid gear, and the like out of any suitable material. In a prefened embodiment, the gear 74 is a spiral gear to provide a smooth tooth action at the operational speeds ofthe centrifugal processing system 10. The upper surface 68 of lower gear insert 70 comprises an axially positioned sleeve 72, which receives and holds gear 74. gear 74 is preferably retained within sleeve 72 by the use of at least one and preferably two spring pins 76 and 76' which are positioned within spring pin holes 73 and 73' extending horizontally through lower gear insert 70 into sleeve 72. Thus, when gear 74 having spring pin receptacles 77 and 77' is inserted into sleeve 72 the spring pins 76 and 76' enter the conesponding receptacles 77 and 77' thus holding the gear 74 in place. Of course, other assembly techniques may be used to position and retain gear 74 within the lower gear assembly 66 and such techniques are considered within the breadth of this disclosure. For example, gear 74 may be held in sleeve 72 by a number of other methods, such as, but not limited to being press fit or frictionally fit, or alternatively gear 74 and lower gear insert 70 may be molded from a unitary body. The base 78 of lower gear insert 70 has a slightly larger diameter than upper body

80 of lower gear insert 70 as a result of a slight flare. This slight flare produces shoulder 82 upon which ball bearing 84 is seated. Once assembled lower bearing assembly 66 is received by sleeve bore 56 extending through neck portion 36 of lower case shell 32. A retaining ring 86 is then inserted into the annular space produced by the difference ofthe outer diameter of the lower bearing assembly 66 and the inner diameter of sleeve bore 56 above ball bearings 84. A second retaining ring 87 (shown in Figure 2) is also inserted into the annular space produced by the difference between the outer diameter ofthe lower bearing assembly 66 and the inner diameter of sleeve bore 56 below ball bearing 84, thereby securing lower gear insert 70 within lower case shell 32. Consequently, ball bearings 62 and 84 are secured by retaining rings 86 and 87, respectively, resulting in lower case shell 32 being joumaled for rotation about lower bearing assembly 66 but fixed against longitudinal and transverse movement thereon. Therefore, when assembled lower bearing assembly 66 is mounted to stationary base 12, by securing screws 14 into threaded holes 79 located in the base 78. Lower case shell 32 is thus able to freely rotate about stationary lower bearing assembly 66 when the drive belt 26 is engaged.

Refening now to Figure 5, extending from the opposite end of neck portion 36 on lower case shell 32 are a number of protrusions or fingers 88, 90, 92, and 94. Positioned between profrusions 88 and 90, and between protrusions 92 and 94 are recessed slots 96 and 98, respectively, for receiving tube guide 102 (Figure 9). The function of tube guide 102 will be discussed in further detail below, but in short it guides umbilical cable 228 connected to cenfrifuge bag 226 through the mid-shaft gear assembly 108 and out ofthe centiifiige 20.

Positioned between profrusions 90 and 92, and between profrusions 88 and 94 are recessed slots 104 and 106, respectively, for receiving gears 110 and 110' of mid-shaft gear assembly 108 (Figure 2). The gears 110 and 110' are preferably configured to provide mating contact with the gear 74 and to produce a desired, overall gear train ratio within the centrifuge 20. In this regard, the gears 110 and 110' are preferably selected to have a similar configuration (e.g., size, tooth number, and the like) as the gear 74, such as a spiral gear design. As illusfrated in Figures 2 and 14 mid-shaft gear assembly 108 comprises a pair of gears 110 and 110'engaged with gears 74 and 131. While the construction of gears and gear combinations is well known to one skilled in the mechanical arts, a brief description is disclosed briefly herein.

Figure 10 illustrates an exploded view depicting the assembly of gear 110, and Figure 11 is a perspective view ofthe gear 110 of Figure 10 as it appears assembled. Gear 110' is consfructed in the same manner. Gear 111 is locked onto mid-gear shaft 112 using key stock 114 and external retaining ring 116. Ball bearing 118 is then attached to mid gear shaft 112 using a flat washer 120 and cap screw 122. Recessed slots 104 and 106 of lower case shell 32 then receive ball bearing 118 and 118' (not shown). In an alternate embodiment ball bearing 118 can be replaced by bushings (not shown). When assembled, gears 110 and 110' make contact with the lower gear 74 (see Figures 2 and 14) to provide contact surfaces for transfening a force from the stationary gear 74 to the gears 110 and

110' to cause the gears 110 and 110' to rotate at a predetermined rate that creates a desired output rotation rate for the driven rotor assembly 200. The rotor assembly 200 is driven by the drive shaft assembly 28 which is rotated by the drive motor 24 at an input rotation rate or speed, and in a prefened embodiment, the drive shaft assembly 28 through the use ofthe gears 110 and 110' is configured to rotate the rotor assembly 200 at an output rotation rate that is twice the input rotation rate (i.e., the ratio ofthe output rotation rate to the input rotation rate is 2:1). This ratio is achieved in the illusfrated embodiment by locking the gears 110 and 110' located within the drive shaft assembly 28 to rotate about the cenfrifuge center axis, A, with the lower case shell 32 which is rotated by the drive motor 24. The gears 110 and 110' also contact the stationary gear 74 which forces the gears 110, 110' to rotate about their rotation axes which are traverse to the centiifiige center axis, A, and as illustrated, the rotation axes ofthe gears 110, 110' coincide. By rotating with the lower case shell 32 and rotating about the gear rotation axes, the gears 110, 110' are able to provide the desired input to output rotation rate of 2:1 to the rotor assembly 200. In this regard, gears 110 and 110' and tabe guide 102 are locked into position by attaching top bearing assembly 124 to lower case assembly 30. Top bearing assembly 124 (as shown in Figure 12) comprises top case shell 126, ball bearing 128, and an upper bearing 130. Top case shell 126, as best seen in Figures 12 and 13, comprises an upper surface 132, a lower lip 134 and a cenfral or axial bore 136 there through. Upper surface

132 slightly overhangs axial bore 136 resulting in a shoulder 138 having a lower surface 140 (shown in Figure 13). Lower lip 134 is a reverse image of upper lip 100 on lower case shell 32 ( shown in Figure 5).

Upper bearing assembly 130 (Figure 12) comprises an upper surface 133 and a lower surface 135 wherein the upper surface 133 has a means for receiving a rotor 202.

On the lower surfare 135 a concentrically positioned column 137 protrudes radially outward perpendicular to lower surface 135. Upper bearing assembly 130 further comprises an axially positioned bore 139 that traverses column 137 and upper surface 133 and receives upper gear insert 131. Upper gear insert 131 also contains an axial bore 142 and thus when positioned concentrically within column 137 axial bores 139 and 142 allow for umbilical cable 228 to travel through upper bearing assembly 130 of top case shell 126 down to cable guide 102 (shown in Figure 14). As discussed previously with respect to lower bearing assembly 66, upper gear insert 131 may be any suitable gear design for receiving an input rotation rate from a mating or contacting gear, such as the gears 110, 110' of the mid-shaft gear assembly 108, with a size and tooth number selected to provide a desired gear train or speed ratio when combined with contacting gears. For example, gear insert 131 may be configured as a straight or spiral bevel gear, a helical gear, a woπn gear, a hypoid gear, and the like. In a prefened embodiment, gear 131 is a spiral gear to provide a smooth tooth action at the operational speeds of the centriftigal processing system 10. Gear insert 131 is preferably retained within column 137 by use of at least one and preferably two spring pins (not shown); however, other assembly techniques may be used to position and retain the gear insert 131 within the column 137 and such techniques are considered within the breadth of this disclosure. For example, gear insert 131 may be held in column 137 by a number of other methods, such as, but not limited to being press fit or frictionally fit or alternatively gear insert 131 and the upper bearing assembly may be molded from a unitary body. Upper bearing assembly 130 is then inserted into axial bore 136 of top case shell 126 so that the lower surface 135 sits flush with upper surface 132 of top case shell 126. Ball bearing 128 is then inserted into the annular space created between the outer diameter of column 137 and the inner side wall 141 of top case shell 126 thereby securing upper bearing assembly 130 into place.

Refening now to Figure 13, lower lip 134 is contoured to mate with protrusions 88, 90, 92 and 94 extending from lower case shell 32. Specifically, the outer diameter of lower lip 134 matches the outer diameter ofthe upper end of main body 40 of lower case shell 32 and recesses 144 and 148 receive and retain protrusions 88 and 92 respectively, while recesses 146 and 150 receive and retain protrusions 94 and S8, respectively. Holes are placed through each recess and each protrusion so that when assembled, fasteners 152 (shown in Figure 12) can be inserted through the holes thereby fastening the top bearing assembly 124 to the lower case assembly 30. Positioned between recesses 144 and

146 and between recesses 148 and 150 are recessed slots 104' and 106', respectively, foi^¬ receiving gears 110 and 110' of mid-shaft gear assembly 108 (Figure 2 and 14). The gears 110 and 110' are preferably configured to provide mating contact with the gear insert 131 and to produce a desired, overall gear train ratio within the cenfrifuge 20. In this regard, the gears 110 and 110' are preferably selected to have a similar configuration (e.g., size, tooth number, and the like) as the gear 131, such as a spiral gear design. Furthermore recessed slots 96' and 98' exist between recesses 144 and 150 and between recesses 146 and 148, respectively. When gears 110 and 110' are assembled as shown in Figure 14, recessed slots 96 and 96' from the lower case shell 32 and top case shell 126, respectively, fom port 154, and recessed slots 98 and 98' form port 156 thereby allowing the umbilical cable 228 to exit centrifuge 20 through either port 154 or 156. Described above is one method of assembling the centrifugal processing system 10 of the present invention; however, those skilled in the art will appreciate that the lower case assembly 30 and upper bearing assembly can be joined in number of ways that allow the four gears to be properly aligned with respect to one another.

In the above manner, the centrifugal processing system 10 provides a compact, portable device useful for separating blood and other fluids in an effective manner without binding or kinking fluid feed lines, cables, and the like entering and exiting the centiifiige 20. The compactness ofthe centrifugal processing system 10 is furthered by the use of the entirely contained and interior gear train described above that comprises, at least in part, gear 74, gears 110 and 110', and gear insert 131 of the upper bearing 130. The gear insert 131 ofthe upper bearing 130 is preferably selected to provide a contact surface(s) with the gears 110 and 110' that transfers the rotation rate ofthe gears 1 10 and 110' and consequently from gear 74 and to the gear insert 131 of the upper bearing 130. In one prefened embodiment, the gear insert 131 ofthe upper bearing 130 is a spiral gear rigidly mounted within the upper bearing 130 to rotate the rotor assembly 200 and having a design similar to that of the spiral gear 74, i.e., same or similar face advance, circular pitch, spiral angle, and the like. During operation, the gear 74 remains stationary as the lower case shell 32 is rotated about the centrifuge axis, A, at an input rotation rate, such as a rotation rate chosen from the range of 0 rpm to 5000 rpm. The gears 110, 110' are rotated both about the centriftige axis, A, with the shell 32 and by contact with the stationaiy gear 74. The spiral gears 1 10, 1 10' contact the gear insert 131 of the upper bearing 130 causing the gear insert 131 and connected upper bearing 130 to rotate at an output rotation rate that differs, i.e., is higher, than the input rotation rate.

Although a number of gear ratios or train ratios (i.e., input rotation rate/output rotation rate) may be utilized to practice the invention, one embodiment of the invention provides for a gear train ratio of 1 :2, where the combination and configuration of the gear 74, gears 110, 110', and gear 131 of the upper bearing 130 are selected to achieve this gear train ratio. Uniquely, the rotation of the gears 110, 1 10' positively affects the achieved gear train ratio to allow, in one embodiment, the use of four similarly designed gears which lowers manufacturing costs while achieving the increase from input to output rotation speeds. Similarly, as will be understood by those skilled in the mechanical arts, numerous combinations of gears in differing number, size, and configuration that provides this ratio (or other selected ratios) may be utilized to practice die invention and such combinations are considered part of this disclosure. For example, although two gears 110, 110' are shown in the mid-shaft gear assembly 108 to distribute transmission forces and provide balance within the operating centrifuge, more (or less) gears may be used to transmit the rotation of gear 74 to the gear of the upper bearing 130. Also, just as the number, size, and configuration of the internal gears may be varied from the exemplary illustration of Figures 1-14, the material used to fabricate the gear 74, the gears 110, 110', and the gear insert 131 may be any suitable gear material known in the art.

Another feature ofthe illusfrated centrifugal processing system 10 that advantageously contributes to compactness is the side-mounted drive motor 24. As illusfrated in Figures 1 and 2, the drive motor 24 is mounted on the stationary base 12 of the enclosure 11 adjacent the centrifuge 20. The drive motor 24 may be selected from a number of motors, such as a standard electric motor, useful for developing a desired rotation rate in the centriftige 20 ofthe centrifugal processing system 10. The drive motor 24 may be manually operated or, as in a preferred embodiment, a motor controller may be provided that can be automatically operated by a confroller ofthe centrifugal processing system 10 to govern operation ofthe drive motor 24 (as will be discussed in detail with reference to the automated embodiment ofthe invention). As illusfrated in Figure 1, a drive belt 26 may be used to rotate the drive shaft assembly 28 (and, therefore, the rotor assembly 200). In this embodiment, the drive belt 26 preferably has internal teeth (although teeth are not required to utilize a drive belt) selected to mate with the external teeth ofthe timing belt ring 46 ofthe lower case assembly 30 portion ofthe drive shaft assembly 28. The invention is not limited to the use of a drive belt 26, which may be replaced with a drive chain, an external gear driven by the motor 24, and any other suitable drive mechanisms. When operated at a particular rotation rate, the drive motor 24 rotates the drive shaft assembly 28 at nearly the same rotation rate (i.e., the input rotation rate). A single speed drive motor 24 may be utilized or in some embodiments, a multi and/or variable speed motor 24 may be provided to provide a range of input rotation rates that may be selected by the operator or by a confroller to obtain a desired output rotation rate (i.e., a rotation rate for the rotor assembly 200 and included centrifuge bag 226. The present invention generally includes an apparatus and methods for the separation of a predetermined fraction(s) from a fluid medium utilizing the principles of centrifugation. Although the principles ofthe present invention may be utilized in a plurality of applications, one embodiment of this invention comprises isolating predetermined fraction(s) (e.g., platelet rich plasma or platelet poor plasma) from anticoagulated whole blood. The platelet rich plasma may be used, for example, in the preparation of platelet concentrate or gel, and more particularly may be used to prepare autologous platelet gel during surgery using blood drawn from the patient before or during surgery.

The cenfrifuge 20 has been discussed above and demonstrates the compact and portable aspects ofthe present invention. To complete the device ofthe present invention a fluid collection device, also refened to as a bowl or rotor 202 is attached to the upper surface 133 ofthe upper bearing assembly 130 as shown in Figures 1 and 2. Rotor 202 is preferably mounted permanently to upper bearing assembly 130, however, rotor 202 may also be capable of being removed. Rotor 202 comprises a rotor base 204 (shown in Figure 15) having a lower annular groove 212, and a rotor cover 206 having an upper annular groove 214. As shown in Figures 17 and 18 the annular interior chamber 216 of rotor 202 is defined by upper and lower annular grooves 212 and 214. The lower annular 212 receives a cenfrifuge bag 226 for containing the fluid medium to be centrifuged. Cenfrifuge bag 226 is connected to supply and receiving containers 398, 400, respectively, via umbilical cable 228 which is preferably, but not limited to a dual lumen. There may be instances where a certain technique requires multiple outlet or inlet ports and consequently umbilical cable 228 ofthe present invention may comprise multiple lumens. Umbilical cable 228 according to the prefened embodiment comprises inlet lumen 230 and outlet lumen 232 such that a fluid medium may be provided to and removed from the cenfrifuge bag 226 during rotation ofthe cenfrifuge rotor 202. One embodiment of centiifiige rotor 202 is more particularly illustrated in Figures

15, 16, 17 and 18. Figure 15 is a perspective view of rotor base 204, and Figure 16 is a perspective view of rotor cover 206. Figure 17 is a cross-sectional side view of rotor 202 taken along view lines 17 in Figure 1, and Figure 18 is a cross-sectional side view of rotor 202 taken along view lines 18 in Figure 1. As illustrated in Figure 15, rotor base 204 comprises raised annular rim 208 and raised column 218 that is axially disposed in base

204. Raised column 218 further has a groove 222 extending across the diameter of column 218. Annular groove 212 is defined by raised annular rim 208 and raised column 218. The height of rim 208 is equal to the height of column 218. Rotor cover 206 shown in Figure 16 comprises raised annular rim 210 and raised column 220 which is axially disposed in rotor cover 206. Raised colunm 220 further has a groove 224 extending across the diameter of colunm 220. Annular groove 214 is defined by rim 210 and column 220. The height of rim 210 is equal to the height of column 220.

Generally, when cenfrifiige rotor 202 is to be assembled for use, a flexible cenfrifuge bag such as a doughnut-shaped cenfrifiige bag 226 (Figure 19 and 20) having a center core 242 is placed in rotor base 204 such that center colunm 218 extends through the core 242 of cenfrifiige bag 226 and the cenfrifiige bag 226 lies in annular groove 212. Rotor cover 206 is superimposed on rotor base 204 such that grooves 222 and 224 are aligned, as illusfrated in Figures 17 and 18. When rotor cover 204 is secured to rotor base 206 by appropriate screws, fasteners, or the like (not shown), rims 208 and 210 are in complete contact with each other such that annular groove 212 and annular groove 214 define rotor interior chamber 216. In one embodiment, columns 218 and 220 are in complete contact with each other. Alternatively, the inner perimeter 240 of centrifuge bag 226 is secured between columns 218 and 220 such that columns 218 and 220 do not completely physically contact each other. With the above description of one embodiment of the centrifuge in mind, another prefened embodiment of a cenfrifuge for use in the centrifugal processing system 10 will be described. Referring to Figures 19-22, a prefened embodiment of a centrifuge 640 is illusfrated that utilizes a uniquely ananged internal pulley system to obtain a desired input to output drive ratio (such as 2:1, as discussed above) rather than an internal gear assembly. The centrifuge 640 utilizes the side-mounted motor 24 (shown in Figure 1) through drive belt 26 to obtain the desired rotation rate at the rotor portion ofthe cenfrifiige.

Refening first to Figure 19, the centrifuge 640 includes a rotor base 644 (or top plate) with a recessed surface 648 for receiving and supporting a centrifuge bag during the operation ofthe cenfrifuge 640. The rotor base 644 is rigidly mounted with fasteners (e.g., pins, screws, and the like) to a separately rotable portion (i.e., a top pulley 698 discussed with reference to Figures 20 and 21) of a lower case shell 660. A cable port 656 is provided centrally in the rotor base 644 to provide a path for a centrifuge tabe or umbilical cable that is to be fluidically com ected to a cenfrifuge bag positioned on the recessed surface 648 of the rotor base 644. It is important during operation of the centrifuge 640 to minimize and confrol contact and binding ofthe umbilical cable and moving parts (such as drive belts and pulleys). In this regard, the lower case shell 660 includes a side cable port 662 for the umbilical cable to enter the centrifuge 640, which, significantly, the side cable port 662 is located between idler pulleys 666, 668 to provide a spacing between any inserted tabe or cable and the moving drive components ofthe centrifuge 640. Idler shaft or pins 664 are mounted and supported within the lower case shell 660 to allow the pins 664 to physically support the pulleys 666, 668. The idler pulleys 666. 668 are mounted on the pins 664 by bearings to freely rotate about the central axis of the pins 664 during operation ofthe centrifuge 640. The idler pulleys 666, 668 are included to facilitate translation ofthe drive or motive force provided or imparted by the drive belt 26 to the lower case shell 660 to the rotor base 644, as will be discussed in more detail with reference to Figures 20 and 21, and to physically support the internal drive belt 670 within the centrifuge 640. The drive belt 26 is driven by the side-mounted motor 24 (shown in Figure 1) and contacts the lower case shell 660 to force the lower case shell 660 to rotate about its cenfral axis. The lower case shell 660 is in turn mounted on the base 674 in a manner that allows the lower case shell 660 to freely rotate on the base 674 as the drive belt 26 is driven by the side-mounted motor 26. The base 674 is mounted to a stationary base 12 (shown in Figure 1) such that the base 674 is substantially rigid and does not rotate with the lower case shell 660.

Referring now to Figures 20-22, the cenfrifuge 640 is shown with a cutaway view to more readily facilitate the discussion ofthe use ofthe internal pulley assembly to obtain a desired output to input ratio, such as two to one. As shown, the base 674 includes vibration isolators 676 fabricated of a vibration absorbing material such as rubber, plastic, and the like through which the base 674 is mounted relatively rigidly to the stationaiy base 12 (of Figure 1). The drive belt 26 from the side-mounted motor 24 (of Figure 1) contacts (frictionally or with the use of teeth and the like as previously discussed) a drive pulley

680, which is rigidly mounted to the lower case shell 660. As the drive belt 26 is driven by the motor 24, the lower case shell 660 through drive pulley 680 rotates about its center axis (which conesponds to the center axis ofthe centrifuge 640). This rotation rate ofthe lower case shell 660 can be thought of as the input rotation rate or speed. To obtain a desired, higher rotation rate at the rotor base 644, the lower case shell

660 is mounted on the base to freely rotate about the centrifuge center axis with bearings 690 that mate with the base 674. The bearings 690 are held in place between the bottom pulley 692 and the base 674, and the bottom pulley 692 is rigidly attached (with bolts or the like) to the base 674 to remain stationary while the lower case shell 660 rotates. The illusfrated bearings 690 are two piece bearings which allow the lower case shell 660 to rotate on the base 674. An internal drive belt 670 is provided and inserted through the lower case shell 660 to contact the outer surfaces ofthe bottom pulley 692. The belt 670 preferably is installed with an adequate tension to tightly mate with the bottom pulley 692 such that frictional forces cause the belt 670 to rotate around the stationary bottom pulley 692. This frictional mating can be enhanced using standard rubber belts or belts with teeth (and of course, other drive devices such as chains and the like may be substituted for the belt 670).

The internal drive belt 670 passes temporarily outside the cenfrifiige 640 to contact the outer surfaces of the idler pulleys 666 and 668. These pulleys 666, 668 do not impart further motion to the belt 670 but rotate freely on pins 664. The idler pulleys 666, 668 are included to allow the rotation about the cenfrifuge center axis by lower case shell 660 to be translated to another pulley (i.e., top pulley 698) that rotates about the same axis. To this end, the idler pulleys 666, 668 provide non-rigid (or rotable) support that assists in allowing the belt 670 to be twisted without binding and then fed back into an upper portion of the lower case assembly 660 (as shown clearly in Figures 20 and 21). As the internal drive belt 670 is fed into the lower case assembly 660, the belt 670 contacts the outer surfaces of a top pulley 698.

During operation ofthe centrifuge 640, the movement of the internal drive belt 670 causes the top pulley 698 to rotate about the cenfrifuge center axis. The idler pulleys 666 and 668 by the nature of their placement and orientation within the cenfrifuge 640 relative to the pulleys 692 and 698 cause the rotor base 644 to rotate in the same direction as the lower case shell 660. Significantly, the top pulley 698 rotated about the cenfrifiige center axis at twice the input rotation rate because it is mounted to the lower case shell 660 via bearings 694 (preferably, a two piece bearing similar to bearings 690 but other bearing configurations can be used) which are mounted to the center shaft 686 of the lower case shell 660 to frictionally contact an inner surface ofthe top pulley 698. Since the internal drive belt 670 is rotating about the bottom pulley 692 and the idler pulleys 666, 668 are rotating about the cenfrifiige central axis by drive belt 26, the top pulley 698 is turned about the centrifuge cenfral axis in the same direction as the lower case shell 660 but at twice the rate.

In other words, the drive force ofthe drive belt 26 and the internal drive belt 670 are combined by the components ofthe cenfrifiige 640 to create the output rotation rate.

While a number of output to input drive ratios may be utilized, as discussed previously, a 2:1 ratio is generally preferable, and the centrifuge 640 is preferably configured such that the second, faster rotation rate ofthe top pulley 698 is substantially twice that ofthe lower case shell 660. The use of an internal drive belt 670 in combination with two pulleys rotating about the same axis and the structural support for the pulleys within a rotating housing results in a centrifuge that is very compact and that operates effectively at a 2:1 drive ratio with relatively low noise levels (which is desirable in many medical settings). The 2:1 drive ratio obtained in the top pulley 698 is in turn passed on to the rotor base 644 by rigidly attaching the rotor base 644 to the top pulley 698 with fasteners 652. Hence, a cenfrifuge bag placed on the recessed surface 648 ofthe rotor base 644 is rotated at a rate twice that ofthe umbilical cable 228 that is fed into lower case shell 660, which effectively controls binding as discussed above. The bearing 694 (one or more pieces) wrap around the entire center shaft 686 ofthe lower case shell 660. To provide a path for the umbilical cord 228 to pass through the centrifuge 640 to the rotor base 644 (which during operation will be enclosed with a rotor top or cover as shown in Figure 1), the rotor base 644 includes the cable port 656 and the center shaft 686 is configured to be hollow to foπn a center cable guide. This allows an umbilical cable 228 to be fed basically parallel to the centrifuge center axis to the centrifuge bag (not shown). The lower case shell 660 includes the side cable port 662 to provide for initial access to the centrifuge 640 and also includes the side cable guide (or tunnel) 684 to guide the cable 228 through the lower case shell 660 to the hollow portion of the center shaft 686. The side port 662 and the side cable guide 684 are positioned substantially centrally between the two idler pulleys 666, 668 to position the cable 228 a distance away from the internal drive belt 670 to minimize potential binding and wear. The cenfrifuge 640 illusfrated in Figures 19-22 utilizes two piece bearings for both the bottom and top pulleys 692 and 698, respectively, and to provide a path for the umbilical cable 228 a central "blind" pathway (via side cable guide 684, the hollow center ofthe center shaft 686, and cable ports 656, 662) was provided in the centrifuge 640. While effective, this "blind" pathway can in practice present binding problems as the relatively stiff cable 228 is fed or pushed through the pathway. To address this issue, an alternate cenfrifiige embodiment 700 is provided and illustrated in Figures 23 and 24. In this embodiment, the upper portions ofthe centrifuge 700 include a guide slot between the idler pulleys 666, 668 that enables an umbilical cable 228 to be fed into the centrifuge 700 from the top with the no components to block the view ofthe operator inserting the cable 228. To allow a guide slot to be provided, the contiguous upper bearing 694 in the cenfrifiige 640 are replaced with bearing members that have at least one gap or separation that is at least slightly larger than the outer diameter ofthe cable 228. A number of bearing members may be utilized to provide this cable entry gap and are included in the breadth of this disclosure. As illustrated, the cenfrifiige 700 includes a rotor base 702 that is rigidly fastened with fasteners 704 to the top pulley 698 (not shown) to rotate with this pulley at the output rate (e.g., twice the input rate) and to receive and support a cenfrifuge bag on recessed surface 716. The rotor base 702 further includes the cable port 718 which is useful for aligning the center ofthe bag and cable 228 with the center ofthe cenfrifuge 700. To allow ready insertion ofthe cable 228 in the centrifuge 700, the rotor base 702 further includes a cable guide slot 712 which as illustrated is a groove or opening in the rotor base 702 that allows the cable 228 to be inserted downward through the cenfrifuge 700 toward the side cable guide 724 ofthe lower case shell 720. The lower case shell 720 also includes a cable guide slot 722 cut through to the top ofthe side cable guide 724. Again, the guide slots 712 and 724 are both located in a portion ofthe cenfrifiige 700 that is between the idler pulleys 666, 668 to position an inserted cable 228 from contacting and binding with the internal drive belt 670, which basically wraps around 180 degrees ofthe top pulley or lower case shell 720.

As shoλvn in Figure 23, the bearing members 706 are spaced apart and preferably, at least one of these spaces or gaps is large enough to pass through the cable 228 to the center shaft ofthe lower case shell 720. As illusfrated, four cam followers are utilized for the bearing members 706, although a different number may be employed. The cam followers 706 are connected to the top pulley to enable the top pulley to rotate and are connected, also, to the center shaft ofthe lower case shell 720 to rotate with the lower case shell 720. The cam followers 706 ride in a bearing groove 710 cut in the lower case shell 720. To provide an unobstructed path for the cable 228, the cable guide slots 712 and 722 are positioned between the two cam followers 706 adjacent the idler pulleys 666, 668, and preferably the guide slots 712, 722 are positioned substantially centrally between the pulleys 666, 668. The guide slots 712, 722 are positioned between these cam followers 706 to position the cable 228 on the opposite side ofthe cenfrifuge 700 as the contact surfaces between the internal drive belt 670 and the top pulley 698 (shown in Figure 20-

22). In this manner, the use of separated bearing members 706 in combination with a pair of cable guide slots 712, 722 allows an operator to readily install the umbilical cable 228 without having to blindly go tlirough the inside ofthe drive system and minimizes binding or other insertion difficulties. A. Flexible, disposable centrifuge bag

One embodiment of disposable, flexible centrifuge bag 226 is more particularly illusfrated in Figures 25 and 26. . The bag is an integral two stage self balancing disposable design. The disposable centrifuge bag 226 has a substantially flat, toroidal- or doughnut-shaped configuration having outer and inner perimeters 238 and 240, respectively, and comprises radially extending upper and lower sheets 234, 236 formed from a substantially flexible material. The upper and lower sheets 234, 236 are superimposed and completely sealed together at outer perimeter 238 by a heat weld, rf (radio frequency) weld or other comparable method of adhering two surfaces. Inner perimeter 240 defines core 242 of bag 226. In one embodiment ofthe invention, cenfrifiige bag 226 further comprises an inlet tube 248 sandwiched between upper and lower sheets 234, 236 and extending from the center of core 242 defined by inner perimeter 240 to the outer perimeter 238 and an outlet tube 250 sandwiched between upper and lower sheets 234, 236 and extending from the center ofthe core 242 to the outer perimeter 238. When upper and lower sheets 234, 236 are sealed together at inner perimeter 240, inlet and outlet tabes 248, 250 are thereby sealed therebetween. Inlet and outlet tubes 248, 250 are each in fluid communication with the interior of centrifuge bag 226 and the environment outside centrifuge bag 226. The length of outlet tube 250 is shorter than the length of inlet tabe 248.

In one embodiment of this invention, outlet tabe 250 is a straight tube as shown in Figure 31. Alternatively, outlet tabe 250 includes a bent fitting 252 fluidly connected to the distal end of outlet tabe 250 (Figures 25 and 26). The bent fitting 252 may be of any number of configurations, although preferably bent fitting 252 is shaped in the form of a "T", "curved T", a "J", or an "L", as illustrated in Figures 27, 28, 29 and 30, respectively. Alternatively, outlet tube 250 and bent fitting 252 may be one contiguous molded unit rather than two connected pieces. Preferably, bent fitting 252 is in the shape of a "T" or a "curved T" as illusfrated in Figures 27 and 28, respectively. The "T" or "curved T" design of bent fitting 252 ensures that the desired blood component (fraction) will be removed from the sides ofthe bent fitting 252, rather than from a fraction located above or below the bent fitting, as discussed below in detail.

When the centiifiige bag 226 is positioned in the annular groove 212 ofthe centrifuge rotor 202 as described above, it is critical that inlet and outlet tubes 248, 250 are seated in groove 222. Further, when rotor cover 206 is positioned over and removably secured to the centrifuge base 204, it is important that inlet and outlet tabes 248, 250 are also seated in groove 224. Seating inlet and outlet tubes 248, 250 in grooves 222, 224 ensures that cenfrifiige rotor 202 is held in a fixed position between rotor base 204 and rotor cover 206 such that the centrifuge bag 226 and cenfrifiige rotor 202 rotate together.

That is, the fixed position of centiifiige bag 226 ensures that centrifuge bag 226 will not rotate independently of centrifuge bag 226 during centrifugation.

Inlet and outlet tabes 248, 250 are fluidly connected at their proximal ends to umbilical cable 228, which in this particular embodiment is a dual lumen tubing connecting cenfrifiige bag 226 to source and receiving containers 398, 400, respectively, for the introduction and removal of components from the centrifuge bag 226 during centrifugation (see Figure 17). Dual lumen tubing 228 comprises inlet lumen 230, which connects inlet tabe 248 of centrifuge bag 226 with source container 398, and outlet lumen 232, which connects outlet tube 250 centrifuge bag 226 with receiving container 400. In one embodiment, the inlet and outlet tabes 248, 250 are adapted at their proximal ends for inserting into the inlet and outlet lumens 230 and 232, respectively. Alternatively, connecting means 254 are inserted into the proximal ends of inlet and outlet tubes 248, 250 for connecting the tubes to the inlet and outlet lumens 230, 232 as illusfrated in Figure 26.

In operation, one end of umbilical cable 228 must be secured to rotor assembly 200 to prevent itself from becoming twisted during rotation of rotor assembly 200 by the coaxial half-speed rotation of drive shaft assembly 28, which imparts a like rotation with respect to the rotor 202 axis and consequently to the umbilical cable 228 that is directed through cable guide 102. That is, if rotor assembly 200 is considered as having completed a first rotation of 360° and drive shaft assembly 28 as having completed a 180° half- rotation in the same direction, the umbilical cable 228 will be subjected to a 180° twist in one direction about its axis. Continued rotation of rotor assembly 200 in the same direction for an additional 360° and drive shaft assembly 28 for an additional 180° in the same direction will result in umbilical cable 228 being twisted 180° in the opposite direction, returning umbilical cable 228 to its original untwisted condition. Thus, umbilical cable 228 is subjected to a continuous flexture or bending during operation of the centrifugal processing system 10 of the present invention but is never completely rotated or twisted about its own axis.

An alternative embodiment of a disposable centrifuge bag of this invention, shown in Figures 35 comprises two or more inlet tubes and/or two or more outlet tubes, wherein the tubes are fluidly connected to a multiple lumen tubing.

The disposable centrifuge bag 226 is formed from a transparent, substantially flexible material, including but not limited to, polyvinyl chloride, polyethylene, polyurethane, ethylene vinyl acetate and combinations ofthe above or other flexible materials. Based upon the flexibility ofthe centrifuge bag 226, the profile ofthe flexible cenfrifuge bag 226, shown in Figures 25 and 26, is determined at least in part by the amount of fluid contained therein. The profile of centrifuge bag 226 is further defined by the interior configuration ofthe cenfrifiige rotor, as discussed below in detail. The ability to manipulate the profile of centrifuge bag 226 based on the interior configuration ofthe centrifuge rotor is utilized at least in part to maximize the volume of fluid medium that can be contained in centrifuge bag 226 during centrifugation, as will be discussed below. The fluid or medium to be centrifuged may be contained within source container 300. For example, when the centrifuge 20 of this invention is used to prepare an autologous platelet gel, the fluid (i.e., whole blood), may be withdrawn from the patient during or prior to surgeiy into source container 398 containing an anticoagulant. The anticoagulated whole blood is introduced to centrifuge bag 226 through inlet tube 248 via inlet lumen 230 after the centrifuge bag 226 has been positioned in the cenfrifuge rotor 202 and rotation thereof is initiated. As discussed above, securing cenfrifuge bag 226 in cenfrifuge base 204 in grooves 222, 224 holds the centrifuge bag 226 in a fixed position therebetween, such that the centrifuge bag 226 cannot move independently ofthe cenfrifiige rotor 202, and therefore the cenfrifiige bag 226 and rotor assembly 200 rotate concunently at the same rate of rotation. Rotation ofthe centrifuge rotor 202 directs the heavier density constituents of the anticoagulated whole blood within the centriftige bag 226 toward the outer perimeter 238 ofthe bag 226, while the lighter density constituents remain closer to an inner region, as illustrated in Figure 32. More specifically, as illusfrated in Figure 32, when the fluid medium being separated is whole blood, the whole blood is separated within centrifuge bag 226 into a red blood cell fraction (256), a white blood cell fraction (258), a platelet rich plasma fraction (260), and a platelet poor plasma fraction (262). As will be appreciated by those of skill in the art, whole blood fractions, red blood cell's and plasma are differently colored, and consequently the separation ofthe fractions can be easily detected by the operator. At an appropriate time during cenhifuging, suction or other drawing means may be applied to the interior of centriftige bag 226 via outlet lumen 232 to remove the desired fraction from the cenfrifiige bag 226. In a further embodiment, centrifuge cover 206 may further contain concentric index lines to assist the operator in viewing the positions of outlet tube 250 to the RBC plasma interface. Based on the speeds and times the location ofthe WBC and platelets can be varied with respect to the red blood cell's and plasma interface. For example, ifthe rpm is held low (approximately 1,000 - 1,700, preferably 1,500) the plasma and platelets will separate from the RBC layer, as the rpm's are increased (1,400 - 1,700) die platelets will separate out ofthe plasma and reside at the plasma to RBC interface in greater concentrations. With increased speeds WBC reside deeper into the RBC pack. With further regard to bent fittings 252, in one embodiment a bent fitting is fluidly connected to the distal end of outlet tube 250. While bent fitting 252 is shown in Figure 32 as having a "T" shape (Figure 27), this is for illustrative purposes only. Thus, it will be appreciated that bent fitting 252 as shown in Figure 32 could have a number of other configurations, such as those shown in Figures 25-31. The design of bent fitting 252 ensures that the desired component is withdrawn (e.g., the platelet rich plasma fraction 260) with less risk of contamination from withdrawing a portion ofthe adjacent fraction 258. Thus, in one embodiment, the desired fraction is withdrawn when its position overlaps with the position of bent fitting 252. Alternatively, the inlet tabe 248 may be first used to draw off the red blood cell fraction 256, and when it is desirable to remove the predetermined fraction from the centriftige bag 226, the predetermined fraction is drawn through bent fitting 252 and outlet tube 250 and directed to receiving container 400 via outlet lumen 232.

With continued reference to Figure 32, as the separation ofthe fluid medium is initiated by centrifiigation, substantially annular regions having constituents of a particular density or range of densities begin to form. For purposes of illustration, the separation of whole blood will be discussed, and as shown in Figure 32 four regions are represented, each of which contains a particular type of constituent of a given density or range of densities. Moreover, it should be appreciated that there may be a given distribution of densities across each ofthe regions such that the regions may not be sharply defined.

Consequently, in practice the regions may be wider (e.g., a larger radial extent) and encompass a range of densities of constitaents.

In the example of Figure 32, the first region 256 is the outermost ofthe four regions and contains red blood cells. The second region 258 contains white blood cells, which have a lower density than that ofthe red blood cells. The third region 260 contains the platelet rich plasma fraction, and the innermost region 262 contains the least dense platelet poor plasma fraction. In one embodiment, it may be desired to harvest the platelet rich plasma fraction in region 260. In order to remove the platelet rich plasma fraction from the centrifuge bag 226, vacuum or suction is provided via outlet lumen 232 to the centrifuge bag 226 to remove a desired portion of region 260. A portion of the fraction 260 that is in the area ofthe bent fitting 252 is drawn through bent fitting 252 and into an appropriate one ofthe collection containers 400 (Figure 17).

More specifically, Figures 33-39 illustrate one method of this invention for the separation of whole blood components, which is a dynamic process. Figure 33 shows one portion ofthe centrifuge bag 226, illustrating the separation ofthe whole blood components after infusion of an aliquot of whole blood into centriftige bag 226 and centrifugation for approximately 60 seconds to 10 minutes at a rate of rotation between 0 and 5,000 rpms. It will be understood by those of skill in the art that faster speeds of rotation will separate the blood in a shorter prior of time. Figure 33 shows the four separated whole blood fractions, with the denser fractions closer to outer perimeter 238, and the less dense fractions closer to inner perimeter 240. While it is well-known that hematocrits (i.e., the volume of blood, expressed as a percentage, that consists of red blood cells) will vaiy among individuals, ranging from approximately 29% - 68%, such variations are easily adjusted for as a result ofthe novel design of centrifuge bag 226 and consequently will not affect the isolation of any ofthe desired fractions as discussed below in detail. Thus, for illusfrative purposes, it will be assumed that centrifugation of an initial infusion of an aliquot of anticoagulated whole blood will give the profile shown in Figure 33. In one embodiment, it is desired to harvest the platelet rich plasma fraction 260. This may be achieved by performing a batch separation process or a continuous separation process as described below.

In one embodiment of a batch separation process of this invention for harvesting the platelet rich plasma fraction 260, cenfrifuge bag 226 has a design as shown in Figure 32 wherein bent fitting 252 positioned approximately in the area where a platelet rich plasma fraction 260 is typically found after centrifugation of an aliquot of whole blood. This approximation is simplified by the placement of concentric indicator lines 205, 207, and 209, (not shown) in the upper surface of rotor cover 206, wherein the concentric lines 205, 207 and 209 conespond approximately with the edges of regions 260, 258, and 256, respectively. Alternative, concentric lines similar to 205, 207 and 209 may be directly imprinted onto the surface of centrifuge bag 226. After centrifugation of an aliquot of blood contained in centrifuge bag 226, a substantial portion ofthe platelet rich plasma fraction 260 is withdrawn from cenfrifiige bag 226 through bent fitting 252 while cenfrifiige rotor 202 is still spinning. As the volume ofthe platelet rich plasma fraction 260 is reduced upon withdrawal, the innermost fraction 262 naturally moves in the direction ofthe outer perimeter 238 due to centrifugal force, as shown in Figure 34. The withdrawal of platelet rich plasma fraction 260 is terminated at a point where the platelet poor plasma fraction 262 is close to bent fitting

252 and before any significant portion of platelet poor plasma fraction 262 could be withdrawn through bent fitting 252, as shown in Figure 34. This point can be detennined either visually by the operator by volume, or by a sensor, as described below in detail. After withdrawal ofthe desired platelet rich plasma fraction 260, inlet lumen 230 is disconnected from the whole blood source container 398 and coimected to a disposal container, after which the remaining fluid in centrifuge bag 226 is evacuated through inlet tabe 248 and directed to the disposal container. The inlet lumen is then reconnected to the whole blood source container, and the above-described batch process is repeated as many times as required until the necessary quantity ofthe desired fraction is isolated. Alternatively, the above-described process can be performed as a continuous process wherein the step of disconnecting the inlet lumen 230 from the whole blood source 398 can be avoided. The continuous process separation of whole blood may be achieve by using a disposable centrifuge bag 226' as illustrated in Figures 39-39 comprising an inlet tube 248 and three outlet tabes 245, 247 and 250, wherein the tubes are connected to an umbilical cable comprising four lumens. More specifically, a disposable centrifuge bag for use in a continuous separation of whole blood comprises inlet tabe 248 connected via an inlet lumen to a whole blood source container, a first outlet tube 250 connected to a first outlet lumen that is in turn connected to a platelet rich plasma receiving container, a second outlet tabe 245 coimected via a second outlet lumen to either a red blood cell receiving container or a waste container and a third outlet tube 247 connected via a third outlet lumen to a platelet poor plasma receiving container. In the continuous separation process, after withdrawal ofthe portion of platelet rich plasma or other cellular components as described above with reference to Figures 33 and 34. Centrifuge bag has the capacity to receive an additional volume (aliquot) of whole blood. Consequently, as shown in Figure 35 infusion of an aliquot of whole blood is reinitiated tlirough first inlet tabe 248 with continued cenhifugation until the capacity ofthe centiifiige bag 226' is reached. As a result ofthe additional volume of blood, the profile of the blood fractions in centrifuge bag 226' will approximately assume the profile shown in Figure 35. As can be seen in Figure 35, the additional volume of blood results in a shift of the location ofthe blood fractions, such that the platelet rich plasma fraction 260 has shifted back into the area ofthe bent fitting 252, and the platelet poor plasma fraction 262 has shifted back towards the inner perimeter 240 and away from the vicinity ofthe bent fitting 252. Additional platelet rich plasma 260 can now be removed from centrifuge bag 226' through outlet tabe 250 as shown in Figure 35.

As described above, removal of an additional volume ofthe platelet rich plasma fraction 260 results in a shift in the location ofthe platelet poor plasma fraction 262 closer to the outer perimeter 238 and consequently closer to the vicinity of bent fitting 252, as shown in Figure 36, at which point removal of platelet rich plasma is again temporarily terminated.

Additional infusions of whole blood aliquots to cenfrifuge bag 262' and removal of platelet rich plasma (by shifting the position of the platelet rich plasma fraction 260 relative to the position ofthe bent fitting 252) as described above may be repeated a number of times. Eventaally, however, the continued infusion of whole blood followed by removal of only the platelet rich plasma fraction will necessarily result in a gradual increase in the volumes (and consequently the widths) ofthe remaining blood fractions 256, 258 and 260 in centrifuge bag 226'. In particular, the volume, and therefore the width, ofthe red blood cell fraction 256 will increase to the extent that the other fractions are pushed closer to the inner perimeter 240 (Figure 37). As shown in Figure 37, the increased volume of red blood cells now present in cenfrifiige bag 226' shifts the location of the fractions towards the inner perimeter 240 such that the white blood cell fraction 260 is now in the vicinity ofthe bent fitting 252 as opposed to the desired platelet rich plasma fraction 262.

The novel design of cenfrifuge bag 226' advantageously provides means for shifting the fractions back to the desired locations when the situation shown in Figure 37 arises. That is, second outlet tube 245 serves as an inlet conduit for introduction of whole blood aliquots into centrifuge bag 226', also serves the function of withdrawing fractions that are located close to the outer perimeter 238. This is achieved in part by attaching the second outlet lumen to either a red blood cell receiving container or a waste container having a suction means (e.g., syringe, pump, etc.) As shown in Figure 38, second outlet tabe 245, having its distal end close to outer perimeter 238, can be operated to withdraw a substantial volume ofthe red blood cell fraction 256, which in turn shifts the location of the remaining fractions 258, 260, 262. The withdrawal ofthe red blood cell fraction 256 may be monitored visually by the operator, or by other means such as a sensor. Alternatively, the positions ofthe fractions may be shifted by withdrawing the platelet poor plasma fraction 262 through third outlet tube 247, which is connected via a third outlet lumen to a platelet poor plasma receiving container. Figure 37 shows that, after withdrawal of a portion ofthe red blood cell fraction

256, the cenfrifuge bag 226' again has the capacity to receive an additional volume of whole blood for centrifugation. An additional infusion of an aliquot of whole blood through inlet tube 248 into the centriftige bag 226' of Figure 37 and centrifugation will produce the profile illusfrated in Figure 39. The above-described steps may be repeated as needed until the desired amount of platelet rich plasma has been harvested. All ofthe above-described steps occur while the cenfrifuge rotor 202 is spinning.

The above-described continuous separation method was illustrated in terms of performing the whole blood infusion step and the platelet rich plasma harvesting step sequentially. An alternative embodiment involves performing the infusion and harvesting steps substantially simultaneously, that is, the platelet rich plasma fraction is withdrawn at approximately the same time as an additional aliquot of whole blood is being added to the bag. This alternate embodiment requires that the centrifuge rotor spin at a rate that results in almost immediate separation ofthe blood components upon infusion of an aliquot of whole blood. As stated previously, all ofthe above-described steps may be monitored either visually by the operator by volume, or by a sensor. Ifthe steps are to be visually monitored, cenfrifiige cover 206 may further include one or more concentric indicator circles 205, 207, 209 (shown in Figures 17 and 18) which may be spaced from the center of cover 206 at distances approximately equal to the outer edges of regions 260, 258 256, respectively, to aid the operator in visualizing the positions of these regions with respect to

252. Figures 33-39 illustrate one embodiment of how the design of centrifuge bags 226 and 226' permit the general locations ofthe various blood fractions to be shifted to allow for continuous harvesting of a desired blood fraction without the risk of contaminating the harvested blood fraction, and further allow for continual on-line harvesting of a large volume (10 to 5 L's) of blood using a small, portable centrifuge device comprising a 10 cc to 200 cc capacity disposable centrifuge bags 226 and 226'.

For example, the design of centrifuge bag 226 having inlet tabe 248 and outlet tube 250 means that the desired component or fraction will be withdrawn from centrifuge bag 226 only through outlet tube 250, while the addition of whole blood aliquots or the removal of other components (e.g., red blood cell fraction 256) will proceed only through dual functional inlet tube 248. In this respect, the harvested fraction (e.g., platelet rich plasma fraction 260) is never withdrawn through inlet tube 248 which was previously exposed to other fluid media (e.g., whole blood or red blood cells). Thus, the design of cenfrifuge bag 226 offers a significant advantage over conventional centrifuge containers comprising only one tube which serves to both infroduce the fluid medium to the container and to withdraw the harvested fraction from the container.

Furthermore, because of its unique design, the use of centrifuge bags 226 and 226' are independent of composition ofthe whole blood to be centrifuged. For example, as stated above, hematocrits (i.e., the percent volume of blood occupied by red blood cells) vary from individual to individual, and consequently the profile illustrated in Figure 32 will vary from individual to individual. That is, the width of red blood cell fraction 256 may be wider or narrower, which in tarn will result in the platelet rich plasma fraction 260 being positioned further away in either direction from bent fitting 252. However, as discussed above in detail with particular reference to Figures 33-34, the design of cenfrifuge bags 226 and 226' allow the location ofthe desired fraction to be shifted until it is in the region of bent fitting 252. Such shifting can be brought about, for example using cenfrifiige bag 226, by withdrawing the red blood cell fraction through inlet tube 248, or by adding whole blood aliquots through inlet tube 248.

An alternative embodiment of a disposable, flexible cenfrifuge bag 270 is illustrated in Figure 40. The disposable centrifuge bag 270 has a substantially flat, toroidal- or doughnut-shaped configuration having outer and inner perimeters 271 and 272, respectively, and comprises radially extending upper and lower sheets 273, 274 fonned from a substantially flexible material. The upper and lower sheets 273, 274 are superimposed and completely sealed together at outer perimeter 271 by an rf weld, heat weld or other comparable method of adhering two surfaces. Inner perimeter 272 defines core 275 of cenfrifuge bag 270. In one embodiment ofthe invention, centriftige bag 270 further comprises inlet tube 276 sandwiched between upper and lower sheets 273, 274 and radially extending from the center of core 275 to the outer perimeter 271, and outlet tube 278 sandwiched between upper and lower sheets 273, 274 and extending across the diameter of core 275 and having first and second distal ends 280, 281. When upper and lower sheets 273, 274 are sealed together at inner perimeter 272, inlet and outlet tubes

276, 278 are thereby sealed therebetween. Inlet and outlet tabes 276, 278 are each in fluid communication with the interior of cenfrifuge bag 270 and the environment outside cenfrifiige bag 270. Inlet tabe 276 and outlet tube 278 are fluidly connected to umbilical cable 228 (not shown), which in this particular embodiment is a dual lumen tubing. Inlet tabe 276 is fluidly connected at its proximal end to umbilical cable 22S, preferably by an

L-shaped connector (not shown), and outlet tube 278 is fluidly connected at its center to umbilical cable 228 via a T-shaped connector (not shown).

The disposable cenfrifuge bag 270 is formed from a transparent, substantially flexible material, including but not limited to, polyvinyl chloride, polyethylene, polyurethane, ethylene vinyl acetate and combinations ofthe above or other flexible materials.

Upper and lower sheets 273, 274 of centrifuge bag 270 are further sealed at two portions between the outer perimeter and the inner perimeter. That is, cenfrifuge bag 270 fiirther comprises a first C-shaped seal 282 located between the outer and inner perimeters 271, 272 and having an first concave indentation or well 283 on the concave side of C- shaped seal 282, and a second C-shaped seal 284 located between the outer and inner perimeters 271, 272 and having an second concave indentation or well 285 on the concave side of C-shaped seal 284. First and second C-shaped seals 282 and 284 are formed by sealing portions of upper and lower sheets 273, 274 together by methods known in the art for sealing two surfaces, including but not limited to rf or heat welding. Ends 288 and 289 of first C-shaped seal 282 are bent inward towards the inner core 275, and likewise ends 290 and 291 of second C-shaped seal 284 are bent inward towards the inner core 275. First and second C-shaped seals 282, 284 have their concave sides facing each other such that the first and second indentations 283, 285 are diametrically opposed to each other. That is, when centrifuge bag 270 is viewed from the top as in Figure 40, first and second C-shaped seals 282, 284 are minor images of each other. First and second C-shaped seals

282, 284 together define an outer chamber 292 between the outer perimeter 271 and first and second C-shaped seals 282, 284, wherein the outer chamber 292 has a toroidal configuration and serves as a first processing compartment. First and second C-shaped seals 282, 284 together further define an inner chamber 293 between first and second C- shaped seals 282, 284 and inner perimeter 272, wherein the inner chamber 293 has a toroidal configuration and serves as a second processing compartment. The first and second C-shaped seals 282, 284 are positioned such that ends 288 and 290 are directly opposite and spaced apart from each other to define a first channel 286 therebetween, and such that ends 289 and 291 are directly opposite and spaced apart from each other to define a second channel 287 therebetween, wherein the first and second channels 286, 287 are diametrically opposed and provide fluid communication between the first processing compartment 292 and the second processing compartment 293. Inlet tabe 276 extends through either channel 286 or chamiel 287, and the first and second distal ends 280, 281 of outlet tabe 278 extend into first and second indentations 283, 285, respectively. Cenfrifiige bag 270 is removably secured between rotor base 204 and rotor cover

206 of rotor 202 in a manner as described above so that centrifuge bag 270 is held in a fixed position relative to rotor base 204 and rotor cover 206 during rotation ofthe cenfrifuge rotor 202. As will be appreciated by those of skill in the art, alternative embodiments of rotor base 204 (Figure 15) and rotor cover 206 (Figure 16) will be required to accommodate the design of centrifuge bag 270. Thus, an alternate embodiment of rotor base 204 comprises raised column 218 comprising first and second grooves which are perpendicular to each other and extend the diameter ofthe raised base column 218, such that when rotor 202 is assembled, inlet tube 276 and outlet tube 278 of centrifuge bag 270 are seated in the first and second grooves, respectively, of raised base column 218. Similarly, an alternate embodiment of cover 206 comprises raised column

220 comprising first and second grooves which are perpendicular to each other and extend the diameter ofthe raised cover column 220, such that when rotor 202 is assembled, inlet tabe 276 and outlet tabe 278 are further seated in the first and second grooves, respectively, of raised cover column 220.

As stated above, inlet and outlet tabes 276, 278 are fluidly coimected to umbilical cable 228, which in this particular embodiment is a dual lumen tubing connecting centrifuge bag 270 to source and receiving containers 398, 400, respectively, for the introduction of the fluid to be centrifuged in bag 270 and for the removal of one or more ofthe separated components from the centrifuge bag 270 during rotation ofthe cenfrifuge 20. Dual lumen tubing 228 comprises inlet lumen 230, which connects inlet tube 276 with source container 398, and outlet lumen 232, which connects outlet tube 278 with receiving container 400.

The fluid or medium to be centrifuged using cenfrifiige bag 270 may be contained within source container 398. For example, when the centrifuge 20 of this invention is used to prepare an autologous platelet gel, the fluid (i.e., whole blood), may be withdrawn from the patient during or prior to surgery into source container 398 containing an anticoagulant. The anticoagulated whole blood is introduced to cenfrifuge bag 270 through inlet tube 276 via inlet lumen 230 after the centrifuge bag 270 has been positioned in the centrifuge rotor 202 and rotation thereof is initiated.

Cenfrifuge bag 270 may be used for the separation and isolation of one or more components dissolved or suspended in a variety of fluid media, including, but not limited to, the separation of cellular components from biological fluids. For example, cenfrifuge bag 270 is usefiil for the concentration and removal of platelets from whole blood. Therefore, the following description ofthe separation of platelets from whole blood using centrifuge bag 270 is merely for purposes of illustration and is not meant to be limiting of the use of bag 270. The separation of a fluid medium such as whole blood in centrifuge bag 270 may be considered to be a two-stage separation process. The first stage of the separation of platelets from whole blood involves separation of a platelet suspension from the red blood cells. The platelet suspension is typically plasma rich in platelets, and it is commonly referred to as platelet-rich plasma (PRP). However, as used herein, the term "platelet suspension" is not limited to PRP in the technical sense, but is intended to encompass any suspension in which platelets are present in concentrations greater than that in whole blood, and can include suspensions that cany other blood components in addition to platelets. The second stage ofthe separation comprises separating platelets from the platelet suspension to produce a platelet concentrate. As used herein, the term "platelet concentrate" is intended to encompass a volume of platelets that results after a "platelet suspension" undergoes a subsequent separation step that reduces the fluid volume ofthe platelet suspension. The platelet concentrate may be a concentrate that is depleted of white blood cells and red blood cells.

With reference to Figure 41, stage one of a whole blood separation process using centrifuge bag 270 begins with the introduction of an aliquot of whole blood into centrifuge bag 270 via inlet tube 276 during rotation ofthe centrifuge 20. As the aliquot of whole blood enters outer chamber 292 of cenfrifuge bag 270, it quickly separates radially under the influence of centrifugal force into various fractions within outer chamber 292 based on the densities ofthe components ofthe whole blood, including an outermost fraction containing the red blood cells which pack along the outer perimeter 271 of centrifuge bag 270, and an inner fraction comprising the platelet suspension. The platelet suspension after centrifiigation ofthe first aliquot of whole blood is represented in Figure 41 by ring 294. Continued infusion of whole blood into the first processing compartment 292 adds an additional volume of red blood cells and consequently pushes the platelet suspension inward as represented by ring 295. Additional infusions of whole blood will continue to push the platelet suspension further inward, as represented by rings

296 and 297 until the first processing compartment 292 is substantially filled with red blood cells (the remainder ofthe volume being plasma) such that the platelet suspension is pushed through channels 286 and 287 into second processing compartment 293. As discussed above, the ends 288, 289 and 290, 291 of C-shaped seals 282, 284, respectively, bend inward, which both helps to fimnel the platelet suspension through channels 286, 287 and to minimize the amount of red blood cells that pass through channels 286, 287. The point at which the red blood cells are near the enhance of channels 286, 287 may be monitored either visually or by a sensor, as described below in detail. At this point the infusion of additional aliquots of whole blood is terminated, and the second stage ofthe two-stage separation process begins. During stage two ofthe separation process, the platelet suspension which was pushed through channels 286, 287 into the second processing compartment 293 flow under the influence of centrifugal force towards positions within the second processing compartment 293 that have the greatest radial distances, that is, towards concave wells 283, 285, where the platelets, being the higher density component ofthe platelet suspension, begin to collect and pack. The platelets can then be withdrawn from concave wells 283, 285 through outlet tabe 278. In the above-described two-stage process for the separation of a platelet suspension from whole blood, the first and second C-shaped seals 282, 284 thus serve as physical barriers between the red blood cells and the platelets to facilitate the separation and collection of platelets from whole blood. First and second concave wells 283, 285 act as reservoirs for containing the platelets as they are separated from the platelet suspension in the second stage ofthe separation process.

After withdraλval ofthe platelets from the wells 283 and 285, inlet lumen 230 is disconnected from the whole blood source container, after which the remaining components in cenfrifiige bag 270 are evacuated through inlet tube 276 by applying suction to inlet lumen 230 and are directed to a disposable container. The inlet lumen 230 is then reconnected to the whole blood source container, and the above-described batch process is repeated as many times as required until the desired quantity of platelets has been harvested. An alternative embodiment of a disposable, flexible cenfrifiige bag having inner C- shaped seals is illusfrated in Figure 42 as centrifuge bag 320. Disposable centrifuge bag 320 has a substantially flat, toroidal- or doughnut-shaped configuration having outer and inner perimeters 322 and 324, respectively, and comprises radially extending upper and lower sheets 323, 325 formed from a substantially flexible material. The upper and lower sheets 323, 325 are superimposed and completely sealed together at outer perimeter 322 by an rf weld, heat weld or other comparable method of adhering two surfaces. Inner perimeter 324 defines core 327 of bag 320.

Upper and lower sheets 323, 325 of centrifuge bag 320 are further sealed at two portions between the outer perimeter and the inner perimeter. That is, centrifuge bag 320 further comprises a first C-shaped seal 326 located between the inner and outer perimeters

322, 324, and a second C-shaped seal 328 located between the inner and outer perimeters 322, 324. The first and second C-shaped seals 326, 328 have their concave sides facing each other such that when centriftige bag 320 is viewed from the top as in Figure 36, first and second C-shaped seals 326 and 328 are mirror images of each other. First and second C-shaped seals 326, 328 together define an outer compartment 348 between the outer perimeter 322 and first and second C-shaped seals 326 and 328, wherein the outer compartment 348 has a toroidal configuration. First and second C-shaped seals further define an compartment 350 between first and second C-shaped seals 326, 328 and inner perimeter 324, wherein the inner compartment 350 has a doughnut shaped configuration. The ends 330 and 332 of first C-shaped seal 326 are slightly curved inward towards the inner core 327, and likewise ends 334 and 336 of second C-shaped seal 328 are slightly curved inward towards the inner core 327. The first and second C-shaped seals 326 and 328 are positioned such that ends 330 and 334 of first and second C-shaped seals 326, 328, respectively, are directly opposite and spaced apart from each other, tiiereby defining first channel 335 therebetween, and such that ends 332 and 336 of first and second seals 326 and 328, respectively, are directly opposite and spaced apart from each other, thereby defining second chaimel 337 therebetween, wherein the first and second channels 335 and 337 are diametrically opposed. First and second channels 335 and 337 provide fluid communication between the outer and inner compartments.

Cenfrifuge bag 320 further comprises an inlet port 340, in the lower sheet 325 for introducing fluid into outer compartment 348. Preferably the inlet port 340 is spaced 90 degrees from chaimel 335 however it could also be positioned at an angle greater or less than 90 degrees from chamiel 335. Cenfrifuge bag 320 further comprises first and second outlet ports 344, 346 in lower sheet 325 and positioned within channels 335 and 337 for withdrawing a fluid compartment from centrifuge bag 320. In a prefened embodiment, cenfrifiige bag 320 comprises inlet tabe 338 secured to the outside surface of upper sheet 323 or lower sheet 325 and radially extending from the center of core 327 towards the outer perimeter 322, wherein inlet tube 338 is fluidly connected at its distal end to inlet port 340. Inlet port 340 fluidly connects inlet tube 338 with the outer chamber 348 of centrifuge bag 320. Inlet tabe 338 is fluidly coimected at its proximal end to umbilical cable 228, preferably by an L-shaped connector (not shown).

Further, in a prefeπed embodiment centriftige bag 320 comprises outlet tube 342 secured to the outside surface of upper sheet 323 or lower sheet 325 and extending across the diameter of core 327, wherein one end of outlet tabe 342 is fluidly connected to first outlet port 344 and the other end of outlet tabe 342 is fluidly connected to second outlet port 346. Outlet tube is fluidly coimected at its center to umbilical cable 228 via a T-shaped connector (not shown).

In an alternative embodiment of this invention, centrifuge bag 320 comprises inlet tabe 338 sandwiched between upper and lower sheets 323, 325 and extending radially from the center of core 327 towards outer perimeter 322, wherein inlet tabe 328 is fluidly connected at its distal end to inlet port 340, and outlet tube 342 sandwiched between upper and lower sheets 323, 325 and extending across the diameter of core 327, wherein one end of outlet tabe 342 is fluidly connected to outlet port 344 and the other end of outlet tube 342 is fluidly coimected to outlet port 346. When upper and lower sheets 323, 325 are sealed together at inner perimeter 324, inlet and outlet tabes 338, 342 are thereby sealed therebetween. Inlet tube 338 and outlet tube 342 are fluidly connected to umbilical cable 228 (not shown), which in this particular embodiment is a dual lumen tubing.

Centrifuge bag 320 is removably secured between rotor base 204 and rotor cover 206 of rotor 202 in a manner as described above so that cenfrifiige bag 320 is held in a fixed position relative to rotor base 204 and rotor cover 206 during rotation ofthe centrifuge rotor 202. As will be appreciated by those of skill in the art, alternative embodiments of rotor base 204 (Figure 15) and rotor cover 206 (Figure 16) as discussed above with respect to centrifuge bag 270 will be required to accommodate the design of centrifuge bag 370.

Cenfrifiige bag 370 may be used for the separation and isolation of one or more components dissolved or suspended in a variety of fluid media, including, but not limited to, the separation of cellular components from biological fluids. For example, cenfrifuge bag 370 is useful for the concentration and removal of platelets from whole blood. Therefore, the following description ofthe separation of platelets from whole blood using centrifuge bag 320 is merely for puiposes of illustration and is not meant to be limiting of the use of bag 320. The separation of a fluid medium such as whole blood in centriftige bag 320 may be considered to be a one-stage separation process. With reference to Figure

36, centrifugatioii of whole blood begins with the introduction of an aliquot of whole blood into cenfrifiige bag 320 through inlet port 340 via inlet tube 338 during rotation of the cenfrifuge 20. Inlet tabe 338 is fluidly coimected via inlet lumen 230 of umbilical cable 228 to an anticoagulated whole blood source. As the aliquot of whole blood enters the outer chamber 348 of centrifuge bag 320, it quickly separates radially within outer chamber 348 into various fractions based on the densities ofthe components ofthe whole blood, including an outeπnost fraction containing the red blood cells which pack along the outer perimeter 322 of centrifuge bag 320, and inner fractions containing the platelets and plasma. Continued infusion of whole blood adds an additional volume of red blood cells and consequently pushes the fraction containing platelets inward. Additional infiisions of whole blood will continue to push the platelet-containing fraction further inward until the chamber 348 is substantially filled with red blood cells (the remainder ofthe volume being plasma), such that the platelet-containing fraction is pushed into channels 335, 337 and into the vicinity of outlet ports 344, 346. As discussed above, the ends of C-shaped seals 326, 328 curve slightly inward, which both helps to funnel the platelet-containing fraction into channels 335, 337, and to minimize the amount of red blood cells that flow into channels 335, 337. The point at which the red blood cells are near the entrance of channels 335, 337 may be monitored either visually or by a sensor, as described below in detail. As the platelet-containing fraction enters the vicinity of outlet ports 344, 346, the infusion of whole blood is terminated, and suction or other drawing means is applied to outlet tabe 342 to withdraw the platelet-containing fraction through outlet ports 344, 346.

After withdrawal of substantial portion of the platelet rich plasma, inlet lumen 230 is disconnected from the whole blood source container and connected to a disposal container, after which the remaining components in centrifuge bag 320 are evacuated through inlet port 34- by applying suction to inlet tube 276 and are directed to a disposal container. The inlet lumen 230 is then reconnected to the whole blood source container, and the above-described process is repeated as many times as required until the desired quantity of platelets has been haivested. B. Rigid centrifuge container

As can be appreciated, it may be desirable to maximize the surface area of separated fraction to be harvested, since this maximizes the amount ofthe fraction which may be collected without increasing the potential for introducing impurities into the separation (e.g., adjacent, lighter density components may begin moving into the region of the fraction being harvested), and without increasing the size ofthe centriftige to an undesirable degree.

In order to maximize the amount ofthe desired component (e.g., platelet rich plasma, white blood cells, or platelet poor plasma) which may be harvested, one embodiment of a cenfrifuge container of this invention for the separation of components in a fluid medium (e.g., whole blood), shown in Figures 45-52, is designed to position the desired component (e.g., platelet rich plasma) the platelet rich plasma at a region within the fixed centrifuge container or centrifuge bag so that the desired fraction has a maximum horizontal surface area (i.e., width). Thus, another embodiment of this invention comprises a centrifuge container 500 shown in Figure 45. Figure 45 is a side cross- sectional view of a rigid container 500 comprising a rigid, annular body 510 having an axial core 600 that is closed at the top end 610 and opened at the bottom end 620. Rigid container 500 further comprises an interior collection chamber 580 for receiving and holding the fluid medium to be centrifuged and having an outer perimeter 585 and an inner perimeter 590. The side, cross-sectional profile of chamber 580 is generally an off- centered "figure eight" or "dumbbell" shape, as shown in Figure 46. As used herein, "figure eight" or "dumbbell" shaped means that the height of section A is approximately equal to the height of section C, and the heights of sections A and C are greater that the height of section B. Furthermore, as used herein, "off-center" means that the width Wi from the center of section B to outer perimeter 585 is less than the width W₂ from the center of section B to inner perimeter 590 as shown in Figure 45 and 46.

Rigid container 500 further comprises inlet channel 550 extending radially from core 600 to a point near the outer perimeter 585 and is fluidly connected at its distal end with the outer area of chamber 580. Rigid container 500 further comprises outlet chamiel

554 extending radially from core 600 to the more cenfral portion of chamber 580 (i.e., the nanow portion or "neck" ofthe figure eight cross-section) and is fluidly connected at its distal end with chamber 580. While the inlet and outlet channels 550, 554 are shown in Figure 45 as being fluidly coimected to the top end of chamber 580, the present invention also includes embodiments wherein both channels 550, 554 are in fluid communication with the bottom end of chamber 580, or wherein channel 550 is in fluid communication with the top end of chamber 580 and channel 554 is in fluid communication with the bottom end of chamber 580, or vice versa. Inlet and outlet channels 550, 554 are fluidly connected to dual lumen tubing 228 having an inlet lumen 230 and an outlet lumen 232. Rigid container 500 is removably secured to the upper surface 133 of upper bearing assembly 130 with appropriate screws, fasteners or the like (not shown). Inlet lumen 230 may be connected to a source for fluid medium, and outlet lumen 232 may be connected to a suction means for withdrawing the desired fraction from the chamber 580.

The configuration of chamber 580 is specifically designed to maximize the collection of platelet rich plasma by centrifiigation of anticoagulated whole blood. More particularly, the shape of chamber 580 increases the width ofthe platelet rich plasma fraction when viewed from the top and decreases the depth ofthe platelet rich plasma fraction when viewed from the side, thus allowing the withdrawal of a greater amount of platelet rich plasma. This unique design can be better explained by comparing Figures 44 and 47. Figure 44 shows a side profile of a rigid centrifuge container 500 as shown in Figure 43, having a generally oval profile and containing whole blood that has been separated into four fractions by centrifugation. In Figure 44, width W₃ indicates the relative horizontal width ofthe platelet rich plasma fraction to be harvested, and Dj indicates the relative depth ofthe platelet rich plasma fraction. Figure 47 shows a side profile of rigid centrifuge container 580 of this invention having the above-described off- centered figure eight shape and containing whole blood that has been separated into four fractions by centrifugation. In Figure 47, width W indicates the relative horizontal width of the platelet rich plasma fraction 260 to be harvested, and D₂ indicates the relative depth ofthe platelet rich plasma fraction 260. Width W is necessarily wider than width W₃ in Figure 44. Thus it can be easily appreciated that upon withdrawal ofthe platelet rich plasma fraction 260 from the oval shaped container shown in Figure 44, platelet poor plasma fraction 262 will shift closer to the outlet tabe 554 relatively quickly. In contrast the dumbbell shaped profile of chamber 580 shown in Figure 47 significantly increases the width W₄ while decreasing the average depth D₂, and therefore a greater portion ofthe platelet rich plasma fraction 260 can be withdrawn with greater accuracy before the platelet poor plasma fraction 262 reaches the outlet tube 554. In an embodiment where the platelet rich plasma is to be collected one could design chamber 580 as follows. The configuration of chamber 580, that is, the relative heights A, B, and C as shown in Figure 46, will be detennined based on the typical location ofthe platelet rich plasma fraction 260 after centrifugation of whole blood. For example, in a rigid cenfrifuge container 500 as illustrated in Figure 45, having chamber

580 with a 30 ml capacity and a radius of approximately 65 mm measured from its rotational axis to the edge 630, the platelet rich plasma will collect in chamber 580 at a region at a radial position ranging from about 35 to about 60 mm from the axis. In this region ofthe chamber 580, as illustrated in Figure 46, the chamber 580 has a height of about 10 mm such that the horizontal surface area "B" of this region, illusfrated in Figure

46, is about 4 mm". Consequently, it can be appreciated that because ofthe unique configuration of chamber 580, the surface area of the platelet rich plasma fraction 260 as illusfrated in Figure 47 may be maximized without undesirably increasing the overall size of the rigid centrifuge container 500. It will be appreciated by those skilled in the art that various geometric designs may be utilized depending on the fluid medium being centrifuged and the cellular fraction to be collected. The process for harvesting platelets from whole blood using rigid container 500 may be achieved in a manner similar to that described for bag 226

Rigid cenfrifuge container 500 may be made from any number of rigid, transparent materials that are capable of withstanding typical sterilization conditions, includmg but not limited to acrylic resins, polycarbonate, or any clear thermal plastic. Preferably rigid container 500 is made of a cost-effective material that is relatively inexpensive to dispose of. C. Cenfrifuge rotor having a complex interior geometry An alternate embodiment of a centrifuge rotor of this invention for holding flexible cenfrifuge bag 226 is illustrated in Figures 48-52. Generally and refening to Figures 48 and 49, the centrifuge rotor 755 is defined by a rotor base 760 (Figures 48, 50 and 52) having a lower channel 780, and a rotor cover 770 (Figures 49 and 51) having an upper channel 782. The annular interior chamber 784 (Figure 48) of rotor 755 is defined by lower and upper channels 780, 782, and has a generally off-centered figure eight side cross-sectional configuration specifically designed to maximize the collection of platelet rich plasma by centrifugation of anticoagulated whole blood, as discussed below in detail.

As illusfrated in Figures 51 and 52, rotor base 760 comprises raised annular rim 775 and raised column 786 which is axially disposed in the interior of rotor base 760. Raised colunm 786 further has a groove 790 (Figure 52) extending the diameter of column

786. The height of rim 775 is equal to the height of column 786. As illustrated in Figure 51, rotor cover 770 comprises raised annular rim 777 and raised column 788 which is axially disposed in the interior of cover 770. Raised column 788 further has a groove 792 (Figure 52) extending the diameter of colunm 78S. The height of rim 777 is equal to the height of column 788. Rotor base 760 and rotor cover 770 are preferably made from any number of rigid transparent materials including, but not limited to acrylic resins, polycarbonate, or any clear thermal plastic.

When centrifuge rotor 755 is to be assembled for use, flexible, doughnut-shaped cenfrifuge bag 226 having a center core 242 is placed in rotor base 760 such that center column 786 preferably, but not necessarily, extends tlirough the core of cenfrifuge bag

226, and inlet and outlet tabes 248, 250 of bag 226 are seated in groove 790. Rotor cover 770 is superimposed on rotor base 760 such that grooves 790 and 792 are aligned and further so that inlet and outlet tabes 248, 250 are seated in groove 792. In one embodiment, when cover 770 is appropriately secured to base 760 (e.g., with screws, clamps, or the like), rims 775 and 777 are in complete contact with each other, and columns 786 and 788 are preferably in complete contact with each other, thereby creating chamber 784 (Figure 48). Alternatively when cover 770 is secured to base 760 as described, the inner perimeter of bag 226 is secured between columns 786 and 788 such that the columns do not physically contact each other. When the generally flat, flexible centrifuge bag 226 is contained within chamber

784 prior to the infusion of a fluid medium (e.g., whole blood), it will not fill the entire volume of chamber 784 but rather will have a radially extending, flat shape as centrifuge rotor 755 is spinning. However, after a sufficient volume ofthe fluid medium (e.g., whole blood) has been introduced into flexible bag 226 through inlet tube 248 such that bag 226 is substantially completely filled, it will be appreciated that filled centrifuge bag 226 will conform to the shape of chamber 784 and consequently will have a off-centered figure eight shaped cross-section.

The off-centered figure eight configuration of the chamber 784 is of approximately the same configuration as the rigid bag 500. Therefore, for the same reasons, the shape of chamber 784 (and consequently the shape of filled bag 226), will assume an off-centered figure eight shape wherein the width ofthe platelet rich plasma fraction is greatly increased relative to the width of a filled bag having an elliptical cross-sectional shape (see, for example, Figures 46 and 47).

As discussed above, a number of methods may be utilized to gauge the harvesting of the desired fraction (such as, but not limited to, platelet rich plasma) from the centrifuge bag. For instance, the separation of platelet rich plasma fraction may be indicated by visual observation of a concentric ring containing the platelet rich plasma (which will be a less colored fraction) and an outer red-colored concentric ring containing the red blood cells. In this case, when such fraction(s) have been separated, the platelet rich plasma may be withdrawn from centrifuge bag 226 by bent fitting 160 to direct the platelet rich plasma to the appropriate collector.

As an alternative to the foregoing, sensors may be incorporated as discussed in detail below to detect the presence ofthe platelet rich plasma fraction.

Based upon the foregoing, it can be appreciated that the centrifugal processing system 10 and the cenfrifuge rotors and bags of this invention have a plurality of features which are suited to harvesting platelet rich plasma, white blood cells, platelet poor plasma or red blood cells from a patient's whole blood in accordance with each of the aspects of the present invention. For example, as discussed above, hematocrits (the volume of blood occupied by red blood cells, expressed as a percentage) vary from individual to individual. Thus, depending on the amount of red blood cells present in a particular sample, the exact radial location of various blood components within the centiifiige bag after centrifugation will also vary. The cenfrifuge bags of this invention overcome this issue by having an inlet tabe capable of not only introducing whole blood into the centriftige bag, but also capable of withdrawing some of all ofthe red blood cell fraction as needed to shift the location of the fraction to be harvested into the area of the outlet tube. Such features are presented in centriftige bags 226, 270, 320 and 500. Yet another embodiment of the cenfrifuge bags of this invention which overcomes problems with varying hematocrits is centrifuge bag 226' having multiple outlet tabes.

Additionally, the centrifugal processing system 10 effectively provides a closed system which enhances the potential for maintaining a desired degree of sterility associated with the entire procedure since materials can thus be both provided to and removed from the cenfrifuge bag during rotation ofthe centrifuge via, for instance, a dual lumen tubing connected to a fluid source (e.g., anticoagulated whole blood withdrawn from a patient before or during surgery) and collection containers (i.e., for the preparation of a platelet gel), without interrupting the process, and thus without significant exposure of the materials to environmental conditions.

Moreover, the portable size ofthe centrifugal processing system 10 in combination with the above-described features of shifting the separated fractions and maximizing the surface area ofthe harvested fraction allows for increased processing capabilities autologous platelet gel over larger, conventional centrifuges The on-line harvesting capabilities ofthe centrifugal processing system 10 allows for continuous, dynamic separation and collection of platelet rich plasma, white blood cells, red blood cells and platelet poor plasma, by adjusting the input and removal of fluid medium and separated fractions as described above. Further, the orientation ofthe flexible and rigid centrifuge bags of this invention and ofthe contents therein (e.g., being generally radially extending) is not significantly modified in the transformation from separation to harvesting ofthe various constituents. Moreover, vortexing throughout the contents ofthe cenfrifiige bags of this invention is reduced or eliminated since the centriftigal processing system 10 does not have to be decelerated or stopped for addition of fluid medium or removal ofthe various fractions therefrom. Further, the general orientation of the flexible and rigid centrifuge bags of the invention (e.g., substantially horizontal) is maintained during removal ofthe desired whole blood fraction similar to the orientation ofthe centrifuge bags assumed during centrifiigation to fiirther assist in maintaining the degree of separation provided by centrifugation. Consequently, the potential is reduced for disturbing the fractions to the degree where the separation achieved is adversely affected. Although the present invention has been described with regard to the separation of whole blood components, it will be appreciated that the methods and apparatus described herein may be used in the separation components of other fluid media, including, but not limited to whole blood with density gradient media; cellular components, or sub-sets of the four whole blood components previously defined.

While blood separation and materials handling may be manually controlled, as discussed above, a further embodiment of the present invention provides for the automation of at least portions of the separation and material handling processes. Referring to Figure 53, an automated centrifugal processing system 800 is illustrated that is generally configured to provide automated control over the steps of inputting blood, separating desired components, and outputting the separated components. The following discussion of the processing system 800 provides examples of separating platelets in a blood sample, but the processing system 800 provides features that would be useful for separating other components or fractions from blood or other fluids. These other uses for the processing system 800 are considered within the breadth of this disclosure. Similarly, the specific components discussed for use in the processing system 800 are provided for illusfration purposes and not as limitations, with alternative devices bemg readily apparent to those skilled in the medical device arts.

In the embodiment illustrated in Figure 53, the processing system 800 includes a blood source 802 coimected with a fluid line 804 to an inlet pump 810. A valve 806, such as a solenoid-operated valve or a one-way check valve, is provided in the fluid line 804 to allow confrol of flow to and from the blood source 802 during operation ofthe inlet pump 810. The inlet pump 810 is operable to pump blood from the blood source 802 through the fluid line 818 to a cenfrifiige 820. Once all or a select portion of the blood in the blood source 802 have been pumped to a blood reservoir 824 of the centrifuge 820 the mlet pump 810 is tamed off and the blood source 802 isolated with valve 806. The inlet pump 810 may be operated at later times to provide additional blood during the operation of the processing system 800 (such as during or after the removal of a separated component). The centrifuge 20 preferably includes a flexible centrifuge bag, for example 226, 226', 270, or 320, positioned within the rotor 202 for collecting the input blood, or alternatively rotor 202 may be a rigid container having an off centered figure eight shaped chamber, which may collect blood directly as discussed previously. Thus, while the embodiment described below illusfrates a cenfrifuge having bag 226, it is to be understood that the alternative centrifuge bags disclosed herein may be used in a similar manner. The centrifuge 20 as discussed above has an internal mid-shaft gear assembly 108 that provides the motive force to rotate the rotor assembly 200, and particularly the rotor 202, at a rotation rate that is adequate to create centrifugal forces that act to separate the various constitaents or components ofthe blood in the rotor 202. The drive assembly 822 may comprise a number of devices useful for generating the motive force, such as an electric motor with a drive shaft coimected to internal drive components of the cenfrifiige 20. In a prefened embodiment, the^' drive assembly 822 comprises an electric motor that drives a belt attached to an exterior portion ofthe centrifuge 20 and more particularly to the timing belt ring 44. To obtain adequate separation, the rotation rate is typically between about 0 RPM and 5000 RPM, and in one embodiment of the invention, is maintained between about 0 RPM and 5000 RPM. As discussed in detail previously, components of particular densities assume radial positions or belts at differing distances from the cenfral axis A ofthe rotor 202. For example, the heavier red blood cells typically separate in an outer region while lower density platelets separate into a region more proximal to the cenfral axis ofthe rotor 202. Between each of these component regions, there is an interface at which the fluid density measurably changes from a higher to a lower density (i.e., as density is measured from an outer to an inner region), and this density interface is used in some embodiments ofthe centrifugal processing system 10 to identify the location of component regions (as will be discussed in more detail below). In a prefened embodiment, the drive assembly 822 continues to operate to rotate the centrifuge 20 to retain the separation ofthe components throughout the operation ofthe centrifugal processing system 10.

Once blood separation has been achieved within the rotor 202, the outlet pump 830 is operated to pump select components from the rotor 202 through outlet lumen 828. As discussed previously, in relation to the features ofthe disposable blood centrifuge bag 226, the centrifuge bag held within the rotor 202 preferably is configured to allow the selective removal of a separated blood component, such as platelets located in a platelet rich plasma region, by the positioning of an outlet lumen 232 a radial distance from the cenfral axis of the cenfrifiige bag 226. Preferably, this radial distance or radial location for the outlet lumen is selected to coincide with the radial location ofthe desired, separated component or the anticipated location of the separated component. In this manner, the outlet pump 830 only (or substantially only) removes a particular component (such as platelets into container 400) existing at that radial distance. Once all or a desired quantity ofthe particular component is removed from the centrifuge bag 226, operation of the outlet pump 830 is stopped, and a new separation process can be initiated. Alternatively, in a prefened embodiment, additional blood is pumped into the centrifuge by 226 by further operating the inlet pump 810 after or conc rent with operation ofthe outlet pump 830. A concern with fixing the radial distance or location of the outlet port is that each blood sample may have varying levels or quantities of different components. Thus, upon separation, the radial distance or location of a particular component or component region within the cenfrifiige bag 226 varies, at least slightly, with each different blood sample. Additionally, because of the varying levels of components, the size of the component region also varies and the amount that can be pumped out ofthe centrifuge bag 226 by the outlet pump 830 without inclusion of other components varies with each blood sample. Further, the position of the component region will vaiy in embodiments of the separation system 10 in which additional blood is added after or during the removal of blood by the outlet pump S30. To address the varying location of a particular separated component, the centrifugal processing system 10 preferably is configured to adjust the location of a separated component to substantially align the radial location ofthe separated component with the radial location ofthe outlet port. For example, the centriftigal processing system 10 may be utilized to collect platelets from a blood sample. In this example, the centrifugal processing system 10 preferably includes a red blood cell collector 812 coimected to the inlet pump 810 via fluid line 814 having an isolation valve 816 (e.g., a solenoid-operated valve or one-way check valve). Alternatively, the pump or syringe may also act as the valve. The inlet pump 810 is configured to selectively pump fluids in two directions, to and away from the centrifuge 820 tlirough fluid line 818, and in this regard, may be a reversible-direction peristaltic pump or other two-directional pump. Similarly, although shown schematically with two fluid lines 804 and 814, a single fluid line may be utilized as an inlet and an outlet line to practice the invention.

Operation ofthe inlet pump 810 to remove fluid from the centrifuge bag 226 is useful to align the radial location ofthe desired separated component with the outlet tabe 250 and inlet tube 248 ofthe centrifuge bag 226. When it is desired to align platelets or platelet rich plasma with the outlet tube 250, the inlet tabe 248 connected to lumen 232 and 230, respectively, inlet tabe 248 is preferably at a greater radial distance than the outlet tube 250. When suction is applied to the inlet lumen 230 by inlet pump 810, red blood cells are pumped out ofthe centrifuge bag 226 and into the red blood cell collector 812. As red blood cells are removed, the separated platelets (i.e., the desired component region) move radially outward to a new location within the cenfrifuge bag 226. The mlet pump 810 is operated until the radial distance ofthe separated platelets or platelet region from the cenfr-al axis is increased to coincide with the radial distance or location ofthe outlet tabe 250 ofthe centrifuge bag 226. Once substantial alignment ofthe desired component region and the outlet tabe 250 is achieved, the outlet pump 830 is operated to remove all or a select quantity ofthe components in the aligned component region.

To provide automation features ofthe invention, the centrifugal processing system 10 includes a controller 850 for monitoring and controlling operation of the inlet pump 810, the centrifuge 20, the drive assembly 822, and the outlet pump 830. Numerous confrol devices may be utilized within the centrifugal processing system 10 to effectively monitor and confrol automated operations. In one embodiment, the controller 850 comprises a computer with a cenfr-al processing unit (CPU) with a digital signal processor, memory, an input output (I/O) interface for receiving input and feedback signals and for transmitting control signals, and software or programming applications for processing input signals and generating confrol signals (with or without signal conditioners and/or amplifiers). The controller 850 is communicatively linked to the devices ofthe centriftigal processing system 10 with signal lines 860, 862, 864, 866, and 868 which may include signal conditioning devices and other devices to provide for proper communications between the confroller 850 and the components ofthe centrifugal processing system 10. Once blood is supplied to the blood source container 802, the operator pushes the start button and the confroller 850 transmits a control signal over signal line 864 to the drive assembly 822, which may include a motor controller, to begin rotating the centrifuge 20 to cause the components ofthe blood in cenfrifuge bag 226 to separate into radially- positioned regions (such as platelet rich plasma regions). After initiation ofthe centrifuge spinning or concunently with operation ofthe drive assembly S22, the confroller 850 generates a confrol signal over signal line 860 to the inlet pump 810 to begin pumping blood from the blood source container 802 to the centrifuge bag 226 ofthe cenfrifuge 20. In some embodiments ofthe processing system 800, the drive assembly 822 is operable at more than one speed or over a range of speeds. Additionally, even with a single speed drive shaft the rotation rate achieved at the centrifuge 20 may vary. To address this issue, the processing system 10 may include a velocity detector 858 that at least periodically detects movement of the centrifuge bag 226 portion ofthe cenfrifuge 20 and transmits a feedback signal over signal line 866 to the controller 850. The controller 850 processes the received signal to calculate the rotation rate ofthe centrifuge 20, and if applicable, transmits a control signal to the drive assembly 822 to increase or decrease its operating speed to obtain a desired rotation rate at the centriftige bag 226.

To determine when separation ofthe components in the centrifuge bag 226 is achieved, the processing system 800 may be calibrated to account for variations in the centiifiige 20 and drive assembly 822 configuration to determine a minimum rotation time to obtain a desired level of component separation. In this embodiment, the confroller 850 preferably includes a timer mechanism 856 that operates to measure the period of time that the cenfrifuge 20 has been rotated by the drive assembly 822 (such as by beginning measuring from the transmission ofthe control signal by the confroller 850 to the drive assembly 822). When the measured rotation time equals the calibrated rotation time for a particular cenfrifuge 20 and drive assembly 822 configuration, the timing mechanism 856 informs the controller 850 that separation has been achieved in the centrifuge bag 226. At this point, the controller 850 operates to transmit confrol signal over signal line 860 to the input pump 810 to cease operation and to the outlet pump 830 over signal line 868 to initiate operation to pump a separated component in the component region adjacent the outlet port of lumen 232 of cenfrifiige bag 226 through fluid line 828. In another embodiment where rotation time is utilized by confroller 850, the velocity feedback signal from the velocity detector 858 is utilized by the confroller 850 to adjust the rotation time as necessary to obtain the desired level of component separation. For example, the centriftigal processing system 10 can be calibrated for a number of rotation rates and the conesponding minimum rotation times can be stored in a look up table for retrieval by the confroller 850 based on a calculated rotation rate. Rotational rates may be varied either manually or automatically to optimize cellular component position and or concentration.

Because the location of component separation regions varies during separation operations, a prefened embodiment ofthe centrifugal processing system 800 includes a sensor assembly 840 to monitor the separation of components within the centriftige bag and to transmit feedback signals over line 862 to the controller 850. As will be understood by those skilled in the art, numerous sensor devices exist for detecting the presence of certain components in a fluid, and specifically a blood, sample. Many of these devices comprise a source of radiant energy, such as infrared, laser, or incandescent light, and a compatible radiant energy-sensitive detector that reacts to the received energy by generating an electric signal. Briefly, these radiant energy devices are useful because the detected signal varies in a measurable fashion with variances in the density ofthe material through which beams ofthe radiant energy are passed. According to the invention, the sensor assembly 840 may comprise any of these well-known types of radiant energy source and detector devices and other sensor devices usefiil for measuring the existence of constituents of fluids such as blood. The source and the detector ofthe sensor assembly 840 are preferably located withm the centriftigal processing system 800 to allow monitoring ofthe cenfrifuge bag 226 and, particularly, to identify the presence of a particular blood component in a radial position coinciding with the radial position ofthe outlet port ofthe cenfrifuge bag 226. In one embodiment, the radiation beams from the source are transmitted through a "window" in the cenfrifiige bag 226 that has a radial location that at least partially overlaps the radial location ofthe outlet port. During operation ofthe centrifugal processing system 800, the feedback signals from the detector ofthe sensor assembly 840 allow the controller 850 to identify when a density interface has entered the window. This may occur for a number of reasons. When red blood cells are being removed by operation ofthe inlet pump 810 to remove fluid from the centrifuge bag 226 via the inlet tube 248. The change in density may also occur when a denser component is being added to the centrifuge bag 226 causmg the particular blood component to be pushed radially inward. In the centrifugation of whole blood, this occurs when additional blood is added by operation ofthe input pump 810 and red blood cells collect in a region radially outward from the platelet region. To account for differing movement ofthe density interface, the window ofthe radiation source may be alternatively positioned radially inward from the location ofthe outlet tabe 250 ofthe cenfrifuge bag 226. By positioning the window inward from the outlet tabe 250, the controller 850 can identify when the outlet pump 830 has nearly removed all ofthe particular component ofthe monitored region and/or when the inlet pump 810 has removed a quantity of denser components causing the monitored region to move radially outward. The confroller 850 can then operate to send confrol signals to rum off the outlet pump 830 or the inlet pump 810 (as appropriate) to minimize the amount of undesired components (lower density components) that enter the outlet tube 250. Alternatively, the sensor assembly 840 may have two radiation sources and detectors, and the second window ofthe sensor assembly 840 may be located a distance radially outward from the outlet tabe 250. With two sensing windows, the sensor assembly 840 is operable to provide the confroller 850 information about a density interface moving radially inward toward the outlet tube 250 (such as when red blood cells are added). In response, the controller 850 can generate a confrol signal to the inlet pump 810 to operate to pump the denser components, such as red blood cells, out ofthe cenfrifuge bag 226. Two sensing windows also allow the controller 850 to detect a density interface moving outward, which allows the confroller 850 to shut off the outlet pump 830 (and/or the inlet pump 810 to stop evacuating processes) and/or to start the inlet pump 810 to add additional blood.

To fiirther clarify operation ofthe processing system 800, Figure 54 is provided which illusfrates the timing and relationship of confrol signals generated by the controller 850 and the receipt of feedback signals from the sensor assembly 840. In this embodiment, the radiation detector ofthe sensor assembly 840 is positioned adjacent outlet tabe (inlet to the outlet pump 830) in the centrifuge bag 226 to sense density changes in the fluid flowing past the outlet tube 250. As illusfrated, operation ofthe processing system 800 begins at time to, with the inlet pump 810, the outlet pump 830, and the centrifuge drive assembly 822 all being off or not operating. At time t₁₍ the controller

850 operates in response to operator input or upon sensing the blood source 802 is adequately filled (sensor not shown) to generate a confrol signal on line 864 to begin operating the centrifuge drive assembly 822 to rotate the cenfrifuge bag 226. In some embodiments, this control signal over line 864 also contains rotation rate information to initially set the operating speed ofthe drive assembly 822. Concunently or at a selected delay time, the controller 850 generates a confrol signal on line 860 to start the inlet pump

810 in a configuration to pump fluid to the centrifuge bag 226 over fluid line 818. The sensor assembly 840 provides an initial density feedback signal to the confroller 850 on line 862, which the controller 850 can process to determine an initial or unseparated density adjacent the outlet tube. Alternatively, the confroller 850 may be configured to request a feedback signal from the sensor assembly 840 after a set delay period (as measured by the timer mechanism 856) to allow separation ofthe components being pumped into the cenfrifiige bag 226 (such as the calibrated, minimum rotation time discussed above) into regions.

At time t₂, the confroller 850 functions to align the region having the desired density, such as a region comprising a higher density of platelets, adjacent the detector of the sensor assembly 840 (i.e., adjacent the outlet tabe). To achieve alignment, the confroller 850 transmits a confrol signal over line 860 to the inlet pump 810 to stop pumping fluid to the cenfrifiige bag 226, to reverse pumping directions including shutting valve 806 and opening valve 816, and to begin pumping components having a higher density then the particular, desired component from the cenfrifuge bag 226 to the collector

812. For example, when the centrifugal processing system 10 is operated to separate and collect platelets or platelet rich plasma, the inlet pump 810 at time, t₂, is operated to pump out the red blood cell fraction by applying suction at the inlet tube 248 to the cenfrifuge bag 226. At time t₃, the density ofthe fluid adjacent the outlet tabe 250 begins to change as denser components are removed by the inlet pump 810, and the sensor feedback signal being fransmitted to the confroller 850 changes in magnitude. The sensor feedback signal continues to change in magnitude (either becoming stronger or weaker depending on the particular sensor utilized and the material being collected) until at time , when the confroller 850 processes the feedback signal and deteπnines that the density ofthe adjacent fluids is within a desired range. This transition can also be thought of as detecting when an interface between two regions of differing densities passes by the location ofthe detector ofthe sensor assembly 840.

With the region ofthe desired, separated component aligned with the outlet tabe 250, the controller 850 operates at time , to send a confrol signal over line 860 to stop operations ofthe inlet pump 810. Also, at time , or at any time thereafter, the confroller

850 generates a confrol signal over line 868 to begin operation the outlet pump 830 to apply suction at the outlet tabe 250 ofthe centrifuge bag 226 to remove the desired component, such as the platelet rich plasma fraction, from the centrifuge bag 226. At time t5, the sensor feedback signal again begins to change in magnitude as the density ofthe fluid near the outlet tabe 250 begins to change, such as when platelet poor plasma begins to enter the sampling window ofthe sensor assembly 840. At time t₆, the density ofthe fluid adjacent the outlet tube 250 and, hence, in the sampling window is outside of a desired density range (e.g., the fluid has less than a predetermined percentage of platelets or other desired fluid component). In response, the confroller 850 transmits a control signal on line 868 to halt operations ofthe outlet pump S30. Of course, the controller S50 can be operated to transmit the signal to the outlet pump 830 at any time prior to time t₆, such as at a time after time ts, when the density ofthe adjacent fluid begins to change but prior to time tή or based on volume removed. The confroller 850 can then operate any time after time tβ, to halt operation ofthe centrifuge drive assembly 822. Further, as discussed above, operations ofthe separation centrifugal processing system 800 can be repeated with the inlet pump 810 being operated to add additional fluid, e.g., blood, after time t₆. Alternatively, the inlet pump 810 and the outlet pump 830 may be operated concunently to add an additional volume of blood with a conesponding new amount of the component being collected after time , to extend the period of time between detection of the interface at time ti and the detection of an out of range density at time t₆.

In the above discussion ofthe automated processing system 800, a sensor assembly 840 was shown in Figure 53 schematically, and it was noted that the location of a radiant energy source and a detector may be any location within the processing system 800 useftil for obtaining an accurate measurement of separating blood components within the centriftige bag 226. For example, the source and detector can be both positioned within the cenfrifiige 20 at a location adjacent the centrifuge bag 226. In this embodiment, problems may arise with providing proper signal and power line connections to the source and sensor and with accounting for the rotation ofthe centrifuge and portions ofthe sensor assembly 840. Hence, one prefened embodiment ofthe processing system 800 provides for an externally positioned sensor assembly 840 including source and detector to simplify the structure ofthe centrifuge 20 while still providing effective density determinations of fluids within the blood reservoir.

Figure 55 illusfrates a general side view ofthe relevant components of this external sensor embodiment ofthe centrifugal processing system 800. Generally, the centrifuge 20 comprises a rotor extension portion 8S0 and a drive portion 881, which is connected to the drive assembly 822 (connection not shown). Both the cenfrifuge 20 and the rotor extension portion 880 rotate about a cenfral or rotation axis, c_axi_s, ofthe centrifuge 20. As discussed in more detail with respect to the internal gearing features ofthe cenfrifuge 20, the drive portion 881 spins in a ratio of 2 to 1 (or other suitable ratio) relative to the reseivoir extension portion 880 to confrol twisting of inlet and outlet fluid lines to the rotor extension portion S80. The internal gearing features ofthe cenfrifuge 20 also enable the cenfrifuge 20 to effectively obtain rotation rates that force the separation of components with differing densities while limiting the risk that denser components, such as red blood cells, will become too tightly packed during separation forming a solid, dense material that is more difficult to pump or remove from the centiifiige 20. Referring again to Figure 55, the rotor extension portion 880 is shown located on the upper end ofthe centrifuge 20 and includes a centrifuge bag 226 or other receptacle. Preferably, the rotor extension portion 880 is fabricated from a transparent or partially transparent material, such as any of a number of plastics, to allow sensing of fluid densities. The rotor extension portion 880 extends a distance, d_over, beyond the outer edge ofthe centrifuge 20 as measured radially outward from the cenfral axis, c^. The distance, d_over, is preferably selected such that the desired component, such as the platelet rich plasma fraction, to be collected readily separates into a region at a point within the centiifiige bag 226 that also extends outward from the centrifuge 20. In this regard, the rotor extension portion 880 is also configured so that the centrifuge bag 226 extends within the rotor extension portion 880 to a point near the outer circumference of the rotor extension portion 880. The distance, d_0Ver, selected for extending the rotor extension portion 880 is preferably selected to facilitate alignment process (discussed above) and to confrol the need for operating the input pump 810 to remove denser components. In one embodiment, the distance, d_over, is selected such that during separation of a typical blood sample center ofthe platelet rich region is about one half the extension distance, d_0Ver, from the circumferential edge ofthe cenfrifuge 20.

The sensor assembly 840 is entirely external to the centrifuge 20 as shown in Figure 55. The sensor assembly 840 includes a source 882 for emitting beams 884 of radiant energy into and tlirough the rotor extension portion 880 and the included centiifiige bag 226. Again, as discussed previously, the radiant energy source 882 may be nearly any source of radiant energy (such as incandescent light, a strobe light, an infrared light, laser and the like) useful in a fluid density sensor and the particular type of detector or energy used is not as important as the external location ofthe source 882. The sensor assembly 840 fiirther includes a detector 886 that receives or senses beams 888 that have passed through the centrifuge bag 226 and have impinged upon the detector 886. The detector 886 is selected to be compatible with the source 882 and to transmit a feedback signal in response sensing the energy beams 888. The detector 886 (in combination with the confroller 850 and its processing capacities) is useful for detecting the density of fluids in the centrifuge bag 226 between the source 882 and the detector 886. Particularly, the sensor assembly 840 is useftil for identifying changes in fluid density and interfaces between fluids with differing densities. For example, the interface between a region containing separated red blood cells and a region containing the platelet rich plasma fraction, and the interface between the platelet rich plasma region and a platelet-poor plasma region.

With some source and detector configurations, a sampling window is created rather than a single sampling point (although a single sampling point configuration is useful as part ofthe invention as creating a window defined by a single radial distance). The sampling window is defined by an outer radial distance, douτ, from the central axis, c_aλ,_s and an inner radial distance, di_N. As may be appreciated, for many source and detector configurations the size ofthe sampling window may be rather small approximating a point and may, of course vary in cross-sectional shape (e.g., circular, square, rectangular, and the like). As discussed previously, it is preferable that the sensor assembly 840 be positioned relative to the reservoir extension portion 880 and the centrifuge bag 226 such that the sampling window created by the source 882 and detector 886 at least partially overlaps the radial position ofthe region created during separation processes containing a component of particular density, such as platelets. This may be a calibrated position determined through calibration processes ofthe cenfrifuge 20 in which a number of blood

(or other fluid) samples are fully separated and radial distances to a particular region are measured. The detennined or calibrated position can then be utilized as a initial, fixed location for the sensor assembly 840 with the source 882 and detector 886 being positioned relative to the rotor extension portion 880 such that the sampling window overlaps the anticipated position ofthe selected separation region. Of course, each sample may vary in content of various components which may cause this initial alignment to be inaccurate and operations ofthe centrifugal processing system 800 may cause misalignment or movement of regions. Hence, alignment processes discussed above preferably are utilized in addition to the initial positioning ofthe sampling window created by the sensor assembly 840.

In an alternate embodiment, the sensor assembly 840 is not in a fixed position within the separation system 800 and can be positioned during separation operations. For example, the sensor assembly 840 may be mounted on a base which can be slid radially inward toward the cenfrifuge 20 and radially outward away from the centrifuge 20 to vary the distances, dm and dour- This sliding movement is useful for providing access to the cenfrifiige bag 226, such as to insert and remove a disposable bag. During operation, the sensor assembly 840 would initially be pushed outward from the centrifuge 20 until a new bag was inserted into the centrifuge bag 226. The sensor assembly 840 could then be slid inward (or otherwise moved inward) to a calibrated position. Alternatively, the centrifugal processing system 800 could be operated for a period of time to achieve partial or full separation (based on a timed period or simple visual observation) and then the sensor assembly 840 slid inward to a position that the operator ofthe centrifugal processing system 800 visually approximates as aligning the sampling window with a desired region of separated components (such as the platelet rich plasma region). The effectiveness of such alignment could then readily be verified by operating the sensor assembly 840 to detect the density ofthe fluids in the centrifuge bag 226 and a calculated density (or other information) could be output or displayed by the controller 850. This alternate embodiment provides a readily maintainable centrifugal processing system 800 while providing the benefits of a fixed position sensor assembly 840 and added benefits of allowing easy relative positioning to obtain or at least approximate a desired sample window and separation region alignment.

In some situations, it may be preferable to not have a rotor extension portion 880 or to modify the rotor extension portion 880 and the sensor assembly 840 such that the extension is not significant to monitoring the separation within the blood reseivoir or centrifuge bag 226. Two alternative embodiments or anangements are illusfrated in Figures 56 and 57 that provide the advantages of an external sensor assembly 840 (such as an external radiation source and detector). With these further embodiments provided, numerous other expansions ofthe discussed use of an external sensor will become apparent to those skilled in the arts and are considered within the breadth of this invention. Refening to Figure 56, a rotor 202 is illusfrated that has no extending portion (although some extension may be utilized) and contains the centrifuge bag 226. Again, the rotor 202 and centrifuge bag 226 are preferably fabricated from plastics or other materials that allow radiation to pass through to detect changes in densities or other properties of fluid samples within the centrifuge bag 226. In this embodiment ofthe sensor assembly 840, the radiation source 882 and the detector 886 are not positioned on opposing sides ofthe rotor 202. Instead, a reflector 885 (such as a mi ror and the like) is positioned within the drive portion 881 ofthe centrifuge to receive the radiation beams 884 from the radiation source 882 and direct them through the portion 880 and centrifuge bag 226. The detector 886 is positioned within the sensor assembly 840 and relative to the cenfrifiige 20 to receive the deflected or reflected beams 888 that have passed through the fluid sample in the centriftige bag 226. In this manner, the sampling window within the centriftige bag 226 can be selected to align with the anticipated location ofthe fraction that is to be collected upon separation. In a prefened embodiment, the sampling window at least partially overlaps with the location ofthe outlet tabe ofthe blood reservoir or cenfrifuge bag 226. In one embodiment, the drive portion is fabricated from a non-transparent material and a path for the beams 884 from the radiation source 884 to the reflector 885 is provided. The path in one prefened embodiment is an opening or hole such as port 154 or 156 (Figure 14) in the side of the drive portion 881 that creates a path or tunnel through which the beams 884 travel unimpeded. Of course, the opening may be replaced with a path of fransparent material to allow the beams to travel to the reflector 885 while also providing a protective cover for the internals ofthe drive portion 881. A path is also provided downstream of the reflector 885 to allow the beams 884 to travel through the drive portion 881 internals without or with minimal degradation. Again, the path may be an opening or tunnel tlirough the drive portion leading to the portion 202 or be a path created with transparent materials. The beams 884 in these tunnel path embodiments enter the drive portion 881 one time per revolution ofthe drive portion 881, which provides an acceptable rate of sampling. Alternatively, a reflector 885 may readily be provided that extends circumferentially about the center axis ofthe drive portion 881 to provide a sampling rate equivalent to the rate of beam 884 transmission. Of course, the positions of the radiation source 882 and the detector 886 may be reversed and the angle of the reflector 885 and transmission ofthe beams 884 may be altered from those shown to practice the invention.

A further embodiment of an external sensor assembly 840 is provided in Figure 57. In this embodiment, the radiation source 882 also acts as a radiation detector so there is no need for a separate detector. In this more compact external sensor configuration, the radiation source and detector 882 transmits beams 884 into the rotating drive portion 881 through or over the path in the drive poition 881. The reflector 885 reflects the beams 884 toward the rotor 202 and the centriftige bag 226 to create a sampling window within the cenfrifuge bag 226 in which density changes may be monitored. After passing through the centrifuge bag 226 and included fluid sample, the beams 888 strike a second reflector 887 that is positioned within the rotor 202 to reflect the beams 888 back over the same or substantially the same path through the centrifuge bag 226 to again strike the reflector 885. The reflector 885 directs the beams 888 out ofthe drive portion 881 and back to the radiation source and detector 882 which, in response to the impinging beams 88S, transmits a feedback signal to the controller 850 for further processing. In one embodiment, the beams 884 enter the driving portion 881 once during eveiy revolution of the driving portion 881. The portion 880 is preferably rotating twice for every station of the driving portion 881, as discussed in detail above, and hence, the second reflector 887 is aligned to receive the beams 888 only on every other rotation of the driving portion 881. Alternatively, a pair of reflectors 887 may be positioned in the rotor 202 such that the beams 888 may be received and reflected back through the cenfrifuge bag 226 once for every rotation of the driving portion 881. In yet a further embodiment, the reflector 885 and second reflector 887 may expand partially or fully about the center axis of the centiifiige 20 (with conesponding openings and/or fransparent paths in the driving portion 881) to provide a higher sampling rate.

According to an important feature ofthe invention, temperature control features are provided in an alternate embodiment of the automated processing system invention

900, as illusfrated in Figure 58. Providing temperature controls within the processing system 900 can take many forms such as controlling the temperature of input fluid samples from the blood source 802, monitoring and controlling the temperature of fluids in the cenfrifuge bag 226 to facilitate separation processes, and controlling the operating temperature of temperature sensitive components of the processing system 900. These components include but are not limited to, red blood cells, white blood cells, plasma, platelet rich plasma or any of these components mixed with other drags, proteins or compounds. In a prefened embodiment of the invention, a temperature confrol system is included in the processing system 900 to heat components removed from the centriftige bag 226 by the outlet pump 830 to a desired temperature range. For example, when the processing system 900 is utilized in the creation of autologous platelet gel, a dispenser assembly 902 is included in the processing system 900 and includes chambers or syringes for collecting and processing platelet rich plasma drawn from the centrifuge 20. As part of the gel creation process, it is typically desirable to activate the platelets in the harvested platelet rich plasma fraction prior to the use of the gel (e.g., delivery to a patient). The temperature control system is useful in this regard for raising, and for then maintaining, the temperature ofthe platelets in the dispenser assembly to a predetermined activation temperature range. In one embodiment of the gel creation process, the activation temperature range is 25° C to 50° C and preferably 37° C to 40° C, but it will be understood that differing temperature ranges may readily be utilized to practice the invention depending on the desired activation levels and particular products being processed or created with the processing system 900.

Refeπing to Figure 58, the temperatare confrol system ofthe processing system 900 includes a temperature confroller 904 that is communicatively linked to the controller 850 with feedback signal line 906. The controller 850 may be utilized to initially set operating temperatare ranges (e.g., an activation temperature range) and communicate these settings over feedback signal line 906 to the temperatare controller 904. Alternatively, the temperatare controller 904 may include input/output (I O) devices for accepting the operating temperature ranges from an operator or these ranges may be preset as part of the initial fabrication and assembly of the processing system 900. The temperature confroller 904 may comprise an electronic confrol circuit allowing linear, proportional, or other confrol over temperatures and heater elements and the like. In a prefened embodiment, the temperature confroller 904 includes a microprocessor for calculating sensed temperatures, memory for storing temperatare and control algorithms and programs, and I/O portions for receiving feedback signals from thenno sensors and for generating and transmitting confrol signals to various temperatare confrol devices (e.g., resistive heat elements, fan rotors, and other devices well-known to those skilled in the heating and cooling arts).

As illusfrated, a temperature sensor 908 comprising one or more temperature sensing elements is provided to sense the temperature ofthe dispenser assembly 902 and to provide a conesponding temperature feedback signal to the temperature controller 904 over signal line 910 (such as an electric signal proportional to sensed temperature changes). The temperatare sensor 908 may be any temperatare sensitive device useful for sensing temperature and, in response, generating a feedback signal usefiil by the temperature confroller 904, such as a thermistor, thermocouple, and the like. In a prefened embodiment, the temperatare sensor 908 is positioned within the dispenser assembly 902 to be in heat transferring or heat sensing contact with the syringes or other chambers containing the separated product which is to be activated. In this manner, the temperature controller 904 is able to better monitor whether the temperatare ofthe relevant chambers within the dispenser assembly 902 is within the desired activation temperature range. To maintain the chambers ofthe dispenser assembly 902 within a temperature range, a heater element 913 is included in the temperature confrol system and is selectively operable by the temperature controller 904 such as by operation of a power source based on signals received from the temperature sensor 908. The heater element 913 may comprise any number of devices useful for heating an object such as the chambers of the dispenser assembly 902, such as a fluid heat exchanger with tubing in heat exchange contact with the chambers. In a prefened example, but not as a limitation, electrical resistance-type heaters comprising coils, plates, and the like are utilized as part ofthe heater element 913. Preferably, in this embodiment, the resistive portions ofthe heater element 913 would be formed into a shape that conforms to the shape ofthe exterior portion ofthe chambers ofthe dispenser assembly 902 to provide efficient heat transfer but preferably also allow for insertion and removal ofthe chambers ofthe dispenser assembly 902. During operation ofthe separation system 900, the temperature controller 904 is configured to receive an operating temperature range, to receive and process temperatare feedback signals from the temperature sensor 908, and in response, to selectively operate the heater element 913 to first raise the temperatare ofthe chambers of the dispenser assembly 902 to a temperature within the operating temperature range and to second maintain the sensed temperature within the operating range.

For example, a desired operating range for activating a gel or manipulating other cellular components and their reactions onto themselves or with agents may be provided as a set point temperatare (or desired activation temperature) with a tolerance provided on either side of this set point temperature. The temperature confroller 904, in this example, may operate the heater element 913 to raise the temperature ofthe chambers ofthe dispenser assembly 902 to a temperature above the set point temperatare but below the upper tolerance temperatare at which point the heater element 913 may be shut off by the temperature confroller 904. When the temperatare sensed by the temperature sensor 908 drops below the set point temperature but above the lower tolerance temperature, the temperature controller 904 operates the heater element 913 to again raise the sensed temperature to above the set point temperature but below the upper tolerance temperature. In this manner, the temperatare confroller 904 effectively maintains the temperature ofthe chambers in the dispenser assembly 902 within a desired activation temperature range (which, of course, may be a veiy small range that approximates a single set temperatare). In one embodiment, the temperature controller is or operates as a proportional integral derivative (PID) temperature controller to provide enhanced temperature control with smaller peaks and abrupt changes in the temperatare produced by the heater element 913. Additionally, the temperature controller 904 may include visual indicators (such as LEDs) to indicate when the sensed temperature is within a set operating range and/or audio alarms to indicate when the sensed temperature is outside the set operating range.

In another embodiment, the heater element 913 is configured to operate at more than one setting such that it may be operated throughout operation ofthe processing system 900 and is not shut off. For example, the heater element 913 may have a lower setting designed to maintain the chambers ofthe dispenser assembly 902 at the lower end ofthe operating range (e.g., acceptable activation temperatare range) with higher settings that provide heating that brings the chambers up to higher temperatures within the set operating range. In another embodiment, the heater element 913 is configured to heat up at selectable rates (e.g., change in temperatare per unit of time) to enhance the activation or other processing of separated liquids in the dispenser assembly 902. This feature provides the temperature controller 904 with control over the heating rate provided by the heater element 913.

As discussed previously, the invention provides features that combine to provide a compact separation system that is particularly adapted for onsite or field use in hospitals and similar environments where space is limited. Figure 59 illustrates one prefened anangement ofthe centrifugal processing system 900 of Figure 58 that provides a compact profile or footprint while facilitating the inclusion of a temperature confrol system. An enclosure 916 is included as part ofthe temperatare confrol system to provide structural support and protection for the components ofthe temperatare confrol system. The enclosure 916 may be fabricated from a number of structural materials, such as plastic. The enclosure 916 supports a heater housing 918 that is configured to allow insertion and removal ofthe chambers and other elements ofthe dispenser assembly 902. The heater housing 918 has a wall that contains the heater element 913 (not shown in Figure 59) which is connected via confrol line 914 to the temperature controller 904. The temperature sensor 908 (not shown in Figure 59) is also positioned within the heater housing 918, and as discussed with reference to Figure 58, is positioned relative to the chambers of the dispenser assembly 902 to sense the temperature of the chambers, and the contained fluid, during operation ofthe system 900. A temperatare feedback signal is transmitted by the temperature sensor 908 over line 910 to temperature controller 904, which responds by selectively operating the heater element 913 to maintain the temperature within the heater housing 918 within a selected operating range.

Because the separation system 900 includes temperature sensitive components, such as the controller 850, the temperature confrol system preferably is configured to monitor and confrol the temperature within the enclosure 916. As illustrated, a temperature sensor 920 is included to sense the ambient temperature within the enclosure

916 and to transmit a feedback signal over line 922 to temperature confroller 904. An air inlet 930, such as a louver, is provided in the enclosure 916 to allow air, Am, to be drawn into and through the enclosure 916 to remove heated air and maintain the temperature within the enclosure 916 at an acceptable ambient temperature. To circulate the cooling air, a fan 934 is provided to pull the air, AI_N, into the enclosure 916 and to discharge hotter air, AOUT_J out of the enclosure 916. The fan 934 is selectively operable by the temperatare controller 904 via control signals over line 938. The size or rating of the fan 934 may vary in embodiments of the invention and is preferably selected based on the volume of the enclosure 916, the components positioned within the enclosure 916 (e.g., the quantity of heat generated by the separation system 900 components), the desired ambient temperatare for the enclosure 916, and other cooling design factors.

In an alternate embodiment of the present invention a dispenser 902, as shown in Figure 60, is provided, for manipulating the cellular fraction which has been isolated and collected via outlet lumen 232. In general, the present invention relates to a dispenser 902 which allows for a manual or automated manipulation of a two-phase method for forming an autologous platelet gel 970 composition wherein all of the blood components for the platelet gel 970 are derived from a patient to whom the platelet gel 970 will be applied.

The methods ofthe present invention for preparing an autologous platelet gel 970 composition, discussed in further detail below, are represented in the flow diagrams depicted in Figures 61-63. As discussed previously, the methods ofthe present invention begins by forming anticoagulated whole blood 396 which is achieved by collecting a patient's whole blood 394 in a source container 398 having an anticoagulation agent, such as sodium citrate (citrate) or heparin. Preferably, the whole blood 394 is collected and mixed with a 3.8% solution of sodium cifrate (refened to herein as "cifrate collection medium") specifically in a 9:1 ratio of blood to cifrate collection medium. A 3.8% solution of sodium citrate is prepared by adding 3.8 grams of sodium citrate per 100 ml of water. While a 3.8% sodium citrate collection medium is that which is frequently used to collect and preserve blood, the person skilled in this art will recognize that the ratio of sodium cifrate to whole blood could be in the range of about 10.9 - 12.9% mMOL, final concenfration. First, as discussed in detail previously and depicted in Figure 61, platelet rich plasma 260 and/or platelet poor plasma 262 are formed by centrifuging a quantity of anticoagulated whole blood 396 that was previously drawn from the patient. The platelet rich plasma 260 is first drawn from the centrifuge bag 226 and into collection chamber 400. Collection chamber 400 is preferably a syringe, but any container that will not contact activate the collected fraction is acceptable. The platelet rich plasma 260 can be pumped via outlet pump 830 (Figure 53) into a collection chamber 400 or the desired fraction can be drawn directly into dispenser 902.

In the prefened embodiment, depicted in Figure 62 according to route 951 , the platelet rich plasma 260, in centrifuge bag 226, is divided into two portions and stored in vessels 952 and 960. The first portion is approximately ! to Vi ofthe total volume of platelet rich plasma 260 and is utilized in phase-one to prepare the thrombin, while the second poition of platelet rich plasma 260 is utilized in phase-two vessel 960. Once the platelet rich plasma 260 or alternatively the platelet poor plasma 262 (shown in Figure 61) is obtained, the prefened methods to obtain thrombin and then produce the platelet gel compositions in an expedited manner, that is, in less than three minutes, are detailed diagrammatically in routes 951 or 981, shown in Figures 62 and 63, respectively and discussed in detail below. If, however, a longer clotting time, that is, in a range of two to eight minutes, is desirous the method to obtain the platelet gel composition ofthe present invention can proceed along the routes 971 and 987, which are also detailed diagrammatically in Figures 63 and 63, respectively and discussed in detail below. Phase-one according to the prefened embodiment (Figure 62) begins by restoring the clot-forming process. To accomplish this, an agent (restoration agent) capable of reversing the effects ofthe anticoagulation agent is added back into the first portion ofthe platelet rich plasma 260 stored in vessel 952. Preferably, the restoration agent can be vessel 952 itself or the restoration agent is contained within vessel 952 prior to the introduction of platelet rich plasma 260; however, the restoration agent may also be introduced later. It is furtheπnore preferable that the contact activator be a material such as but not limited to glass wool 953 or silica, aluminum, diatomaceous earth, kaolin, etc., or non-wettable surfaces such as plastic, siliconized glass, etc. Chemical activators, such as kaolin, can also be used to speed up the clotting time; however, their subsequent removal would also be necessary. In the prefeπed embodiment, a plastic syringe is the prefened container used to collect the desired fraction. In the presently prefened embodiment ofthe invention, the reversal ofthe anticoagulant is accomplished using calcium chloride. However, any substance which is known or found to be functionally equivalent to calcium chloride, such as, calcium gluconate or calcium carbonate, in restoring the coagulation activity of citrated blood may be used in the practice ofthe present invention. Thus, although calcium chloride is the presently prefened calcium salt for use in the invention, any calcium salt which functions in a similar manner to calcium chloride may be used in the invention. Similarly, although many blood coagulation reactions are cu ently believed to require calcium ions as cofactors, any substance which is known or subsequently found to be functionally equivalent to calcium in facilitating these coagulation reactions may be used, either individually or in combination with calcium, in the practice ofthe present invention. Ifthe anticoagulation agent used was heparin, then heparinase or any other suitable anticoagulant reversing compound would be used to reverse the effect ofthe anticoagulation agent. The concentration ofthe restoration agent used to reverse the anticoagulation will depend in part, upon the concentration ofthe anticoagulation agent in the platelet rich plasma 260 and the stoichiometiy ofthe chelating and coagulation reactions. However, the concentration of the restoration agent used to reverse the anticoagulation must be sufficient to achieve clot formation. Upon restoration ofthe platelet rich plasma 260 as shown in Figures 62, a clot 954 will naturally form. The resulting clot 954 is then triturated by squeezing the clot 954 through glass wool 953 which serves not only as a contact activator but also as a filter, thus expressing thrombin 955. Alternatively, or in addition a filter 958 having a large micron pore size thereby allowing the removal of clot debris and any activator or solids that are present. Filter 958 is positioned at the outlet 956 of vessel 952. In the prefened embodiment, the thrombin 955 is then mixed with the second portion of platelet rich plasma (PRP) 260 contained within vessel 960 to form the platelet gel composition 970 of the present invention in less than three minutes and in quantities sufficient for clinical use. Other additives can be added to the above-described process to increase the concenfration of tlirombin formed by the intrinsic pathway or the extrinsic pathway.

As discussed in detail above, restoring the clotting cascade function of citrated plasma by addition of calcium chloride and exposure to an activating agent such as glass wool can generate autologous thrombin. The yield of autologous thrombin by this method however, may be low due to incomplete conversion of protlirombm and the inactivation of generated thrombin by fibrin and antithrombin III. The addition of modifying agents, such as epsilon aminocaproic acid, to the plasma may improve the yield by reducing the amount of thrombin neutralization. The greatest improvement in tlirombin yield, however, will be achieved by providing a thromboplastic material upon which the necessary clotting factors will assemble to maximize the rate of prothrombin conversion. The activated platelet membrane provides such a stimulant surface and also enriches the necessary factor V activity by secreting additional factor V during platelet degranulation. The addition of exogenous lipoprotein and/or thromboplastic material to the plasma environment may also serve to maximize the thrombin generation by activation of both intrinsic and extrinsic clotting cascades. Additional amplification of autologous tlirombin generation may also be attained by pretreafrnent of PRP and or PPP to block or remove both antithrombin-III and fibrinogen prior to conversion of prothrombin to thrombin. Such modification may be attained by use of appropriate adsorptive agents, antibodies or precipitating reagents.

In an alternative embodiment, thrombin 950 is mixed with the platelet poor plasma 262 of phase-two thereby forming the autologous platelet gel composition 972 ofthe present invention in less than three minutes. A third embodiment ofthe present invention, route 971, shown in Figure 62, contemplates collecting the original quantity of platelet rich plasma (PRP) 260 derived from the anticoagulated whole blood 396 in a container, having a wettable surface, such as glass. The platelet rich plasma 260 is then recalcified and the platelet gel composition 974 forms. The desired platelet gel composition 974 will require approximately two to eight minutes to form as opposed to less than a three minute formation as was described in the prefened embodiment.

In the fourth embodiment depicted diagrammatically by route 981 in Figure 63, the platelet poor plasma 262, rather then the platelet rich plasma 260, is divided mto two portions, as discussed previously in the prefened embodiment. The first portion, used in phase-one, which is approximately '/₄ to '/_ the original volume is stored in a vessel 952 having a wettable surface, then the restoration agent, preferably calcium chloride, is added directly to the platelet poor plasma 262. Surface activation ofthe restored platelet poor plasma 262 occurs as result ofthe vessel's surface and/or the glass wool 953 or other surface or chemical activators and a clot 962 thus forms. The resulting clot 962 is triturated, as described previously, and the thrombin 955 is collected. Tlirombin 955 is then mixed with the platelet rich plasma 260 of phase-two thereby foiming the platelet gel sealant composition 973.

In the fifth embodiment, tlirombin 955 is mixed with the platelet poor plasma 262 of phase-two thereby forming the platelet gel composition 975 in less than three minutes.

The sixth embodiment follows route 987, shown in Figure 63 wherein the original quantity of platelet poor plasma 262 is collected in a container having a wettable surface, such as glass. The platelet poor plasma 262 is then recalcified and the platelet gel composition fomis. The tensile strength ofthe platelet gel compositions ofthe present invention can be effected by the addition of calcium ions. Consequently, if a stronger bioadhesive sealant composition is desired using the methods discussed above and disclosed in routes 951 and 981, in Figures 62 and 62, respectively, more calcium ions may be added at the time the serum is introduced into the platelet rich plasma 260 or the platelet poor plasma 262. Alternatively, if the method of preparing the platelet gel compositions follows routes 971 and 987, depicted in Figures 62 and 63, respectively, then calcium ions may be introduced directly into the platelet rich plasma 260 or the platelet poor plasma 262 and the platelet gel compositions 974 and 976, respectively, will form.

As discussed in further detail below, the time period necessaiy for the formation of the platelet gel composition ofthe present invention is dependent on the quantity of serum added. A 1 :4, 1 :2 and 3:4 ratio of serum to platelet rich plasma or platelet poor plasma results in the formation ofthe platelet gel composition in approximately 90, 55 and 30 seconds, respectively. Furthermore, due to the fact that thiOmbin is autocatalytic, it is important that the serum be used λvithin five hours of preparation, preferably within two hours and ideally immediately. Alternatively, the serum can be chilled or frozen indefinitely.

The platelet gel compositions of this invention may be used for sealing a surgical wound by applying to the wound a suitable amount platelet rich plasma or platelet poor plasma. Moreover, due to the fact that the platelet gel compositions ofthe present invention have been prepared solely from blood components derived from the patient that is to receive the platelet gel there is a zero probability of introducing a new blood transmitted disease to the patient. The methods ofthe present invention may be further modified so that the fonned platelet gel composition functions not only as a haemostatic agent, but also as an adjunct to wound healing and as a matrix for delivery of drugs and proteins with other biologic activities. For example, it is well known that fibrin glue has a great affinity to bind bone fragments which is useful in bone reconstruction, as in plastic surgeiy or the repair of major bone breaks. Consequently, in keeping with the autologous nature ofthe platelet gel composition of die present invention autologous bone from a patient can be ground or made into powder or the like, and mixed into the platelet rich plasma obtained in phase-two of the methods of the present invention. Autologous thrombin is then mixed in with the platelet rich plasma and bone fragments in an amount sufficient to allow the resulting gel to be applied to the desired locale where it congeals. Other materials that may be utilized are, but not limited to, gelatin collagen, degradable polymers, hyaluronic acid, carbohydrates and starches.

In instances where the desired platelet gel composition ofthe present invention is to further function as a delivery device of drags and proteins with other biologic activities the method ofthe present invention may be modified as follows. Prior to adding the thrombin, obtained in phase-one, to the platelet rich plasma of phase-two a wide variety of drags and proteins with other biologic activities may be added to the platelet rich plasma of phase-two. Examples of the agents to be added to the platelet rich plasma prior to the addition ofthe serum include, but are not limited to, analgesic compounds, such as Lidocaine, antibacterial compounds, including bactericidal and bacteriostatic compounds, antibiotics (e.g., adriamycin, erythromycin, gentimycin, penicillin, tobramycin), antifimgal compounds, anti-inflanmiatories, antiparasitic compounds, antiviral compounds, anticancer compounds, such as paclitaxol enzymes, enzyme inhibitors, glycoproteins, growth factors (e.g. lymphokines, cytokines), hormones, steroids, glucocorticosteroids, immunomodulators, immunoglobulins, minerals, neuroleptics, proteins, peptides, lipoproteins, tamoricidal compounds, tamorstatic compounds, toxins and vitamins (e.g., Vitamin A, Vitamin E, Vitamin B, Vitamin C, Vitamin D, or derivatives thereof). It is also envisioned that selected fragments, portions, derivatives, or analogues of some or all ofthe above may be used. A number of different medical apparatuses and testing methods exist for measuring and deteπnining coagulation and coagulation-related activities of blood. These apparatuses and methods can be used to assist in determining the optimal formulation of activator, that is, thrombin, calcium and plasma necessary to form the platelet gel composition ofthe present invention. Some ofthe more successful techniques of evaluating blood clotting and coagulation are the plunger techniques illusfrated by U.S.

Pat. Nos. 4,599,219 to Cooper et al, 4,752,449 to Jackson et al, and 5,174,961 to Smith, all of which are assigned to the assignee of the present invention, and all of which are incorporated herein by reference.

Automated apparatuses employing the plunger technique for measuring and detecting coagulation and coagulation-related activities generally comprise a plunger sensor cartridge or cartridges and a microprocessor controlled apparatus into which the cartridge is inserted. The apparatus acts upon the cartridge and the blood sample placed therein to induce and detect the coagulation-related event. The cartridge includes a plurality of test cells, each of which is defined by a tube-like member having an upper reaction chamber where a plunger assembly is located and where the analytical test is carried out, and a reagent chamber which contains a reagent or reagents. For an activated clotting time (ACT) test, for example, the reagents include an activation reagent to activate coagulation ofthe blood. A plug member seals the bottom of a reagent chamber. When the test commences, the contents ofthe reagent chamber are forced into the reaction chamber to be mixed with the sample of fluid, usually human blood or its components. An actuator, which is a part ofthe apparatus, lifts the plunger assembly and lowers it, thereby reciprocating the plunger assembly through the pool of fluid in the reaction chamber. The plunger assembly descends by the force of gravity, resisted by a property of the fluid in the reaction chamber, such as its viscosity. When the property ofthe sample changes in a predetermined manner as a result ofthe onset or occurrence of a coagulation-related activity, the descent rate of the plunger assembly there through is changed. Upon a sufficient change in the descent rate, the coagulation-related activity is detected and indicated by the apparatus.

Using the methods discussed above, cartridges were assembled with serum obtained from either platelet rich plasma or platelet poor plasma, and CaCl₂ in the reagent chambers. Clotting time tests were performed by the automated process with either platelet rich plasma (PRP) or platelet poor plasma (PPP) dispersed into the reaction chambers ofthe cartridges. In the first experiment, the results of which are represented in Figure 64, the amount of serum, the type of plasma from which the serum was derived, and the type of plasma the serum was mixed with were tested to determine the shortest clotting times. The ratios of serum to platelet rich plasma or platelet poor plasma that were studied included 1:4, 1:2, and 3:4. In the second set of experiments, the results of which are represented in Figures 66 and 67, the relationship between actual gel time for the platelet gel ofthe present was compared to the clotting time in the cartridge, wherein there is a 0, 30, or 60 minute delay of adding the serum from its generation. The third set of experiments, the results of which are represented in Figures 68 and 69, studied the effect of calcium addition on actual gel time versus clotting time in the cartridge. The final set of experiments, the results of which are represented in Figure 65, studied the effect of adding calcium on clotting times.

Although clotting times varied among donors, comparisons of clotting times for individual donors show significant effects ofthe serum to plasma ratio and the calcium concenfration. For all donors, the shortest clotting times occuned for the 3:4 ratio, with clotting times that were 47% shorter than those for the 1:4 ratio. Although the difference in clotting times for the 3:4 ratio and the 1 :2 ratio was not statistically significant, the clotting times were consistently shorter using the 3:4 ratio for all donors. These results demonstrate that clotting times may be shortened by increasing the seram to platelet rich plasma ratio. Similarly, clotting times were significantly affected by the amount of calcium added, with the shortest clotting times obtained when no calcium was added, suggesting that the seram contained levels, of calcium that were sufficient to recalcify the citrated platelet rich plasma. Preliminary results from the scale-up experiments suggest that experimental clotting times in the cartridges conelate with actual gel times. The invention is further illustrated by the following non-limited examples. All scientific and technical tenns have the meanings as understood by one with ordinaiy skill in the art. The specific examples which follow illusfrate the methods in which the bioadhesive sealant compositions ofthe present invention may be prepared in a clinical setting and are not to be construed as limiting the invention in sphere or scope. The methods may be adapted to variation in order to produce compositions embraced by this invention but not specifically disclosed. Further, variations ofthe methods to produce the same compositions in somewhat different fashion will be evident to one skilled in the art.

EXAMPLES

The examples herein are meant to exemplify the various aspects of canying out the invention and are not intended to limit the invention in any way.

Example 1

Preparation of Bioadhesive Sealant Composition Using Platelet Rich Plasma and Serum

6 cc's of platelet rich plasma are drawn into receiving chamber 961 and 3 cc's per PRP or PPP are drawn into receiving chamber 957 which further contains 0.33 cc's of 10% calcium chloride and glass wool. Clotting ofthe contents will occur in two to eight minutes in receiving chamber 957. The clot is then squeezed tlirough optional filter 958 and the seram, produced therefrom, is added to the platelet rich plasma contained in receiving chamber 961 by either mixing or spraying the two components. The platelet rich plasma and the seram will gel within approximately three minutes.

The application ofthe gel using the syringe-type devices 902 as described above maybe less than desfrable for may applications. Consequently, in an alternate embodiment the inactive blood component and thrombin can be mixed and/or injected into a mold having a desired geometric shape. The mold may be constructed of a material having a wettable surface, such as, but not limited to plastic. In particular, platelet gel ofthe present invention may be used to temporarily fill, cavities such as but not limited to holes left in the gum from tooth extraction and/or holes left in tissue or bone as a result of injury or surgical procedures. The present invention provides a simpler way of introducing platelet gel for specific uses, by providing that the platelet gel be pre-shaped or molded into a beneficial shape prior to being inserted into a cavity. In the case of tooth extraction the platelet gel may be shaped so as to achieve a basic conical shape. Other shapes such as, but not limited to rods, and rectangles are contemplated by this invention. The ability to cause the gel to be more, or less, solid and thus malleable may be achieved during the activation sequence ofthe gel formation. The foregoing description is considered as illusfrative only ofthe principles ofthe invention. Furthermore, since numerous modifications and changes will readily occur to those skilled in the art, it is not desired to limit the invention to the exact consfruction and processes shown as described above. Accordingly, all suitable modifications and equivalents may be resorted to falling within the scope ofthe invention as defined by the claims which follow. The foregoing description is considered as illusfrative only ofthe principles ofthe invention. The words "comprise," "comprising," "include," "including," and "includes" when used in this specification and in the following claims are intended to specify the presence of one or more stated features, integers, components, or steps, but they do not preclude the presence or addition of one or more other features, integers, components, steps, or groups thereof. Furthermore, since a number of modifications and changes will readily will readily occur to those skilled in the art, it is not desired to limit the invention to the exact construction and process shown described above. Accordingly, all suitable modifications and equivalents may be resorted to falling within the scope ofthe invention as defined by the claims which follow.

Claims

CLAIMSThe embodiments ofthe invention in which an exclusive property or privilege is claimed are defined as follows:

1. A flexible, disposable cenfrifiige bag for receiving and holding a fluid sample, comprising: one or more tabes, and upper and lower flexible sheets, said sheets having doughnut shaped configurations, inner perimeters defining a cenfral core of said centriftige bag, and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer perimeters thereby defining the outer perimeter of said cenfrifiige bag, and wherein said tabes are sandwiched between said upper and lower sheets and extend from the cenfral core of said centrifuge bag, whereby when said upper and lower sheets are sealed together at their inner perimeters said tubes are sealed between said upper and lower sheets at the inner perimeter.

2. The centrifuge bag of claim 1, comprising an inlet tabe having a distal end positioned at a first radial distance from the center axis of said cenfrifiige bag and an outlet tube having a distal end positioned at a second radial distance from the center axis of said cenfrifuge bag, said first radial distance being greater than said second radial distance.

3. The cenfrifiige bag of claim 2, wherein said inlet tube and said outlet tube are in fluid communication at their proximal ends with a dual lumen tubing.

4. The cenfrifuge bag of claim 3, wherein said dual lumen tubing comprises an inlet lumen fluidly coimected to said inlet tube.

5. The centrifuge bag of claim 3, wherein said dual lumen tubing comprises an outlet lumen fluidly connected to said outlet tube.

6. The centrifuge bag of claim 2, wherein said outlet tube is fluidly connected

» at its distal end to a bent fitting.

7. The cenfrifuge bag of claim 6, wherein said bent fitting has a configuration selected from the group consisting of a T shape, a cuived T shape, a J shape, or an L shape.

8. The centrifuge bag of claim 2, comprising an inlet tube and two or more outlet tabes.

9. The centriftige bag of claim 8, wherein said tubes are fluidly coimected to a tubing comprising multiple lumen, wherein the number of lumen equals the number of tubes.

10. The cenfrifiige bag of claim 9, wherein one of said outlet tubes is fluidly connected at its distal end to a bent fitting.

11. The centrifuge bag of claim 1 , wherein said fluid comprises two or more components which separate into two or more fractions at varying radial distances within said bag during centrifugation, wherein a predetermined fraction is collected from said bag.

12. The centrifuge bag of claim 11, wherein said fluid is introduced into said bag via said inlet tabe, and said predetemiined fraction is withdrawn from said bag via said outlet tube.

13. The cenfrifiige bag of claim 1 1, wherein said fluid comprises whole blood and said predetemiined fiaction is platelet rich plasma, platelet poor plasma, white blood cells or red blood cells.

14. The centrifuge bag of claim 1, wherein said flexible sheets are polyvinyl chloride, polyethylene, polyurethane, ethylene vinyl acetate or combinations thereof.

15. A method of separating components contained in a fluid and harvesting a predetermined component, said method comprising: a) providing a flexible centrifuge bag comprising: i) at least inlet tube; ii) at least outlet tabe; and iii) upper and lower flexible sheets, said sheets having doughnut shaped configurations, inner perimeters defining a central core of said centrifuge bag, and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer perimeters thereby defining the outer perimeter of said centrifuge bag, and wherein said inlet and outlet tubes are sandwiched between said upper and lower sheets and extend from the central core of said cenfrifuge bag, such that when said upper and lower sheets are sealed together at their inner perimeters said inlet and outlet tubes are sealed between said upper and lower sheets at the inner perimeter; b) providing a centrifuge rotor; c) securing said flexible centrifuge bag in a horizontal position in said cenfrifiige rotor, whereby said cenfrifuge bag is fixably rotatable with said centrifuge rotor; d) rotating said centrifuge rotor; e) providing said fluid to said rotating cenfrifuge bag via said inlet tube, whereby said components of said fluid separate radially into two or more fractions based on their densities; and f) removing a fraction containing said predetermined component from said rotating centrifuge bag via said outlet tube.

16. The method of claim 15, wherein said inlet tabe has a distal end positioned at a first radial distance from the center axis of said cenfrifuge bag and said outlet tube has a distal end positioned at a second radial distance from the center axis of said centrifuge bag, said first radial distance being greater than said second radial distance.

17. The method of claim 16, wherein said inlet and outlet tubes are fluidly connected to a dual lumen tubing having an inlet lumen in fluid communication with the proximal end of said inlet tube and an outlet lumen in fluid communication with the proximal end of said outlet tube.

18. The method of claim 17, wherein the distal end of said outlet tube is in flow communication with said fraction containing said predetermined component.

19. The method of claim 18, wherein said fluid is anticoagulated whole blood and said predetemiined component is platelet rich plasma, platelet poor plasma, white blood cells or red blood cells.

20. The method of claim 19, wherein said inlet lumen is fluidly connected to an anticoagulated whole blood source.

21. The method of claim 17, wherein said dual lumen tubing is prevented from completely rotating about its axis during rotation of said centrifuge.

22. A method of harvesting a predetermined component contained in a fluid, said method comprising: a) providing a flexible centrifuge bag comprising an inlet tube, a first outlet tabe, a second outlet tabe, and a third outlet tube; b) providing a centrifuge rotor; c) securing said flexible centrifuge bag in said cenfrifuge rotor, whereby said centrifuge bag is fixably rotatable with said centrifuge rotor; d) rotating said centrifuge rotor; e) providing an aliquot of said fluid to said cenfrifiige bag via said inlet tube, wherein said fluid sample separates into fractions of similar density components at varying radial distances from the cenfral axis, wherein the distal end of said first outlet tabe is positioned to be in flow communication with a fraction containing said predetermined component; f) withdrawing said fraction containing said predetermined component from said rotating centrifuge bag via said first outlet tabe; and g) terminating said withdrawing when an undesired fraction is determined to be in the vicinity of said outlet tube.

23. The method of claim 22, wherein said flexible centrifuge bag comprises upper and lower flexible sheets, said sheets having doughnut shaped configurations, inner perimeters defining a central core of said centrifuge bag and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer perimeters thereby defining the outer perimeter of said cenfrifuge bag, and wherein said tabes are sandwiched between said upper and lower sheets and extend from the central core of said cenfrifiige bag toward the bag outer perimeter, such that when said upper and lower sheets are sealed together at their inner perimeters said tabes are sealed between said upper and lower sheets at the inner perimeter.

24. The method of claim 22, further comprising h) withdrawing the outermost fractions from said bag via said inlet tabe; and i) repeating steps d) through h) until a desired quantity of said predetemiined component has been harvested.

25. The method of claim 22, wherein said fluid is anticoagulated whole blood and said predetermined component is platelet rich plasma, platelet poor plasma, white blood cells or red blood cells.

26. The method of claim 22, wherein said tubes are fluidly connected at their proximal ends to a multiple lumen tubing.

27. The method of claim 26, wherein said multiple lumen tubing is prevented from completely rotating about its axis during rotation of said centrifuge.

28. The method of claim 22, wherein said rotor comprises a transparent cover having one or more concentric indicator lines for locating said predetermined component.

29. The method of claim 22, wherein said deteπnining is perfoπned visually or by a sensor.

30. An apparatus for separating one or more predetermined components contained in a fluid sample, comprising: a rotatable cenfrifuge rotor; and a flexible centrifuge bag for receiving and holding said sample, said centriftige bag horizontally positionable within and rotatable with said cenfrifiige rotor, said centiifiige bag comprising: at least one inlet tube and at least one outlet tube, and upper and lower flexible sheets, said sheets having doughnut shaped configurations, inner perimeters defining a cenfral core of said cenfrifuge bag, and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer perimeters thereby defining the outer perimeter of said centrifuge bag, and wherein said tubes are sandwiched between said upper and lower sheets and extend radially from the cenfral core of said centrifuge bag toward the bag outer perimeter, whereby when said upper and lower sheets are sealed together at their inner perimeters said tabes are sealed between said upper and lower sheets at the inner perimeter.

31. The apparatus of claim 30, further comprising a multiple lumen tubing fluidly comiected to said tabes, wherein the number of lumen equals the number of tabes.

32. The apparatus of claim 30, wherein said centrifuge bag comprises a inlet tabe for introducing said sample into said bag, said inlet tube having a first length, and a first outlet tube for withdrawing a predetermined component separated from said sample, said first outlet tabe having a second length, wherein said first length is greater than said second length.

33. The apparatus of claim 32, wherein said components separate at varying radial distances within said cenfrifuge bag based on their densities, wherein the distal end of said first outlet tabe is positioned to be in flow communication with a fraction containing said predetemiined component.

34. The apparatus of claim 30, further comprising a bent fitting fluidly coimected at the distal end of at least one of said outlet tubes.

35. The apparatus of claim 34, wherein said bent fitting has a configuration selected from the group consisting of a T shape, a curved T shape, a J shape, or an L shape.

36. The apparatus of claim 31 , wherein said tubing is prevented from completely rotating about its axis during rotation of said centiifiige.

37. The apparatus of claim 30, wherein said fluid is whole blood and said predetemiined component is platelet rich plasma, platelet poor plasma, white blood cells or red blood cells.

38. A flexible centrifuge bag for separating components of a fluid sample, comprising: upper and lower flexible sheets, said sheets having doughnut shaped configurations, inner perimeters defining a central core of said centrifuge bag and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer perimeters thereby defining the outer perimeter of said centrifuge bag; an outer first processing compartment; an inner second processing compartment disposed between said first processing compartment and said core and having diametrically opposed first and second indentations on its inner surface, wherein said first and second processmg compartments are separated by first and second inner seals; two diametrically opposed channels, wherein said channels provide fluid communication between said first processing chamber and said second processing chamber; at least one inlet tube sandwiched between said upper and lower sheets and radially extending from the center of said core for introducing said fluid sample into said first processing compartment; and at least one outlet tube for withdrawing a select component from said second processing compartment sandwiched between said upper and lower sheets and extending across the diameter of said core, wherein one end of said outlet tabe is proximal to said first indentation and the other end of said outlet tabe is proximal to said second indentation, whereby when said upper and lower sheets are sealed together at their inner perimeters said inlet and outlet tubes are sealed between said upper and lower sheets at the inner perimeter.

39. The centrifuge bag of claim 38, wherein said first and second inner seals are C-shaped seals, wherein the concave sides of said first and second seals face each other, and wherein the ends of said seals define said channels.

40. The centrifuge bag of claim 38, wherein said tubes are fluidly connected to a multi lumen tubing.

41. The centrifuge bag of claim 40, wherein said inlet tube is in fluid communication at its proximal end with an inlet lumen of a multi lumen tubing.

42. The cenfrifiige bag of claim 40, wherein said outlet tube is in fluid communication at its axis with an outlet lumen of a multi lumen tubing.

43. The cenfrifuge bag of claim 38, wherein said components separated radially within said centrifuge bag based on their densities.

44. The cenfrifiige bag of claim 43, wherein said fluid sample is anticoagulated whole blood.

45. The centrifuge bag of claim 44, wherein said first processing compartment separates said whole blood into a red blood cell fraction and a platelet suspension.

46. The cenfrifuge bag of claim 45, wherein said second processing compartment separates platelets from said platelet suspension, wherein said platelets or white blood cells collect in said indentations.

47. A flexible centrifuge bag for separating components of a fluid sample, comprising: upper and lower flexible sheets, said sheets having doughnut shaped configurations, inner perimeters defining a cenfral core of said cenfrifuge bag, and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer perimeters thereby defining the outer perimeter of said cenfrifiige bag; a first C-shaped imier seal between said outer perimeter and said inner perimeter and having a first indentation on its concave side; a second C-shaped inner seal between said outer perimeter and said inner perimeter and having a second indentation on its concave side, wherein the concave side of said first inner seal faces the concave side of said second inner seal, and wherein said first and second indentations are diametrically opposed; a first channel defined by a first end of said first inner seal and a first end of said second inner seal; a second chamiel defined by a second end of said first inner seal and a second end of said second inner seal; an inlet tube sandwiched between said upper and lower sheets and extending from the center ofthe core of said cenfrifuge bag; and an outlet tube sandwiched between said upper and lower sheets and extending across the diameter of said core, wherein one end of said outlet tube is proximal to said first indentation and the other end of said outlet tube is proximal to said second indentation, whereby when said upper and lower sheets are sealed together at their inner perimeters said inlet and outlet tubes are sealed between said upper and lower sheets at the inner perimeter.

48. The cenfrifiige bag of claim 47, wherein said outer perimeter together with said first and second inner seals define a first processing compartment.

49. The cenfrifiige bag of claim 47, wherein said inner perimeter together with said first and second inner seals define a second processing compartment.

50. The centrifuge bag of claim 47, wherein the distal end of said inlet tabe is in fluid communication with said first processing compartment.

51. The centrifuge bag of claim 47, wherein said tabes are fluidly connected to a multiple lumen tubing comprising an inlet lumen and an outlet lumen.

52. The centrifuge bag of claim 51 , wherein said inlet tabe is fluidly connected at its proximal end to said inlet lumen.

53. The centrifuge bag of claim 51 , wherein said outlet tabe is fluidly coimected at its axis to said outlet lumen.

54. The centriftige bag of claim 47, wherein said fluid sample is anticoagulated whole blood.

55. A disposable centrifuge bag for harvesting platelets from anticoagulated whole blood, comprising: upper and lower flexible sheets, said sheets having doughnut shaped configurations, hmer perimeters defining a central core of said centrifuge bag and outer peruneters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer and inner perimeters; a first processing compartment for receiving said whole blood and separating said whole blood into a red blood cell fraction and a platelet suspension fraction; a second processing compartment for separating platelets from said platelet suspension fraction; channels for directing said platelet suspension fraction from said first processing compartment into said second processing compartment; an mlet tube for introducing whole blood into said first processing compartment; and an outlet tube for removing said platelets from said second processing compartment.

56. The centrifuge bag of claim 55, wherein said inlet tabe extends radially from said central core and is sealed between the upper and lower sheets at the inner perimeter.

57. The centrifuge bag of claim 55, wherein said second processing chamber comprises diamefrically opposed first and second wells for collecting and holding said platelets.

58. The centrifuge bag of claim 55, wherein said outlet tabe extends across the diameter of said core and is sealed between said upper and lower sheets at the inner perimeter of said bag, wherein one end of said outlet tabe is proximal to said first well and the other end of said outlet tabe is proximal to said second well.

59. The cenfrifuge bag of claim 55, wherein said tabes are fluidly coimected to a multiple lumen tubing comprising an inlet lumen and an outlet lumen.

60. The centrifuge bag of claim 59, wherein said inlet tabe is fluidly connected at its proximal end to said inlet lumen.

61. The cenfrifuge bag of claim 59, wherein said outlet tabe is fluidly coimected at its axis to said outlet lumen.

62. The centrifuge bag of claim 55, wherein said red blood cell fraction and said platelet suspension separate from said whole blood in a path that extends radially from the rotational axis.

63. An apparatus for harvesting platelets from whole blood, comprising: a) a rotatable centrifuge rotor; and b) a flexible cenfrifuge bag comprising: upper and lower flexible sheets, said sheets having doughnut shaped configurations, inner perimeters defining a central core of said centrifuge bag and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer and mer perimeters; a first processing compartment for receiving said whole blood and separating said whole blood into a red blood cell fraction and a platelet suspension fraction; a second processing compartment for separating platelets from said platelet suspension fraction; channels for directing said platelet suspension fraction from said first processing compartment into said second processing compartment; an inlet tube for infroducing whole blood into said first processing compartment; and an outlet tube for removing said platelets from said second processing compartment.

64. The apparatus of claim 63, wherein said inlet tabe extends radially from said cenfral core and is sealed between the upper and lower sheets at the hmer perimeter.

65. The apparatus of claim 63, wherein said second processing chamber comprises diametrically opposed first and second wells for collecting and holding said platelets.

66. The apparatus of claim 63, wherein said outlet tube extends across the diameter of said core and is sealed between said upper and lower sheets at the inner perimeter of said bag, wherein one end of said outlet tabe is proximal to said first well and the other end of said outlet tube is proximal to said second well.

67. The centrifuge bag of claim 63, wherein said tabes are fluidly connected to a multiple lumen tubing comprising an inlet lumen and an outlet lumen.

68. The apparatus of claim 67, wherein said inlet tabe is fluidly connected at its proximal end to said inlet lumen.

69. The apparatus of claim 67, wherein said outlet tabe is fluidly connected at its axis to said outlet lumen.

70. The apparatus of claim 63, wherein said red blood cell fraction and said platelet suspension separate from said whole blood in a path that extends radially from the rotational axis.

71. The apparatus of claim 67, wherein said tubing is prevented from completely rotating about its axis during rotation of said centrifuge rotor.

72. A method for separating components contained in a fluid, comprising a) providing a flexible centrifuge bag comprising: i) upper and lower flexible sheets, said sheets having doughnut shaped configurations, inner perimeters defining a central core of said cenfrifuge bag and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer perimeters tiiereby defining the outer perimeter of said cenfrifiige bag; ii) a first processing compartment; iii) a second processing compartment disposed between said first processing compartment and said core, said second processing compartment having two diametrically opposed indentations on the inner surface; iv) two diamefrically opposed channels, wherein said channels provide fluid conmiunication between said first processing compartment and said second processing compartment; v) an inlet tabe for introducing said fluid into said first processing compartment, said inlet tabe sandwiched between said upper and lower sheets and radially extending from the center of said core; and vi) an outlet tube for withdrawing a select one of said separated components from said second processing compartment, said outlet tabe sandwiched between said upper and lower sheets and extending across the diameter of said core, wherein one end of said outlet tube is proximal to said first indentation and the other end of said outlet tube is proximal to said second indentation, whereby when said upper and lower sheets are sealed together at their inner perimeters said inlet and outlet tubes are sealed between said upper and lower sheets at the inner perimeter; b) providing a centrifuge rotor; c) securing said flexible centrifuge bag in said centrifuge rotor, whereby said centiifiige bag is fixably rotatable with said centrifuge; d) rotating said cenfrifiige rotor; e) providing said fluid to said centrifuge bag via said inlet tube, whereby said components of said fluid separate radially into fractions based on their densities until said select component flows through said channels into said second processing compartment; and f) removing one or more of said components from said second processing compartment via said outlet tabe.

73. The centrifuge bag of claim 72, wherein said tubes are fluidly coimected to a multiple lumen tubing comprising an inlet lumen and an outlet lumen.

74. The method of claim 73, wherein said inlet tabe is fluidly connected at its proximal end to said inlet lumen.

75. The method of claim 73 , wherein said outlet tube is fluidly coimected at its axis to said outlet lumen.

76. The method of claim 72, wherein said removed components are platelet rich plasma, platelet poor plasma, white blood cells or red blood cells.

77. A method for separating components contained in a fluid sample, comprising: a) providing a flexible centrifuge bag for receiving said fluid sample, comprising: i) upper and lower flexible sheets, said sheets having doughnut shaped configurations, inner perimeters defining a central core of said centrifuge bag, and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer perimeters thereby defining the outer perimeter of said centrifuge bag; ii) a first C-shaped inner seal between said outer perimeter and said imier perimeter and having a first indentation on its concave side; iii) a second C-shaped imier seal between said outer perimeter and said inner perimeter and having a second indentation on its concave side, wherein the concave side of said first C-shaped inner seal faces the concave side of said second C- shaped imier seal, and wherein said first and second indentations are diametrically opposed; iv) a first channel defined by a first end of said first inner seal and a first end of said second iimer seal; v) a second chamiel defined by a second end of said first imier seal and a second end of said second inner seal; vi) an inlet tube sandwiched between said upper and lower sheets and extending from the center ofthe core of said centrifuge bag to a point near said outer perimeter; and vii) an outlet tube sandwiched between said upper and lower sheets and extending across the diameter of said core, wherein one end of said outlet tabe is proximal to said first indentation and the other end of said outlet tube is proximal to said second indentation, whereby when said upper and lower sheets are sealed together at their inner perimeters said inlet and outlet tubes are sealed between said upper and lower sheets at the inner perimeter; b) providing a centrifuge rotor; c) securing said flexible cenfrifuge bag in said cenfrifiige rotor, whereby said centrifuge bag is fixably rotatable with said centrifuge rotor; d) rotating said centrifuge rotor; e) providing said fluid to said cenfrifuge bag via said inlet tabe, whereby said components of said fluid separate radially into fractions based on their densities; f) tenninating said providing when a selected one of said components flows tlirough said channels and collects in said indentations; and g) removing one or more of said separated components from said centrifuge bag via second outlet tube.

78. The cenfrifiige bag of claim 77, wherein said tabes are fluidly connected to a multiple lumen tubing comprising an inlet lumen and an outlet lumen.

79. The method of claim 78, wherein said inlet tube is fluidly coimected at its proximal end to said inlet lumen.

80. The method of claim 78, wherein said outlet tabe is fluidly connected at its axis to said outlet lumen.

81. The method of claim 24, wherein said fluid sample is anticoagulated whole blood.

82. A flexible centrifuge bag for separating components of a fluid sample, comprising: upper and lower flexible sheets, said sheets having doughnut shaped configurations, inner perimeters defining a cenfral core of said centrifuge bag, and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer perimeters thereby defining the outer perimeter of said cenfrifuge bag; an outer compartment; an inlet port in said lower sheet and in fluid communication with said outer compartment; an inner compartment disposed between said outer compartment and said core, wherein said imier and outer compartments are separated by first and second inner seals; first and second channels, wherein said channels are diametrically opposed and provide fluid communication between said outer compartment and said inner compartment; first and second outlet ports in said lower sheet and in fluid conmiunication with said first and second channels, respectively; an inlet tube having its distal end in fluid communication with said inlet port and fixed to the outer surface of said lower sheet and radially extending from the center of said core; and an outlet tube fixed to the outer surface of said lower sheet and extending across the diameter of said core, wherein one end of said outlet tabe in fluid communication with said first outlet port and the other end of said outlet tube is in fluid communication with said second outlet port.

83. The cenfrifuge bag of claim 82, wherein said tabes are fluidly connected to a multiple lumen tubing comprising an inlet lumen and an outlet lumen.

84. The cenfrifuge bag of claim 83, wherein said inlet tabe is fluidly connected at its proximal end to said inlet lumen.

85. The cenfrifuge bag of claim 83, wherein said outlet tube is fluidly coimected at its axis to said outlet lumen.

86. The centrifuge bag of claim 83, wherein said fluid sample is whole blood.

87. A flexible centrifuge bag for separating components of a fluid sample, comprising: upper and lower flexible sheets, said sheets having doughnut shaped configurations, inner perimeters defining a cenfral core of said centrifuge bag and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer perimeters thereby defining the outer perimeter of said cenfrifuge bag; a first C-shaped inner seal between said outer perimeter and said inner perimeter; an inlet port in said lower sheet and positioned between said outer perimeter and said first inner seal for infroducing said fluid into said centrifuge bag; a second C-shaped inner seal between said outer perimeter and said inner perimeter, wherein the concave side of said first inner seal faces the concave side of said second inner seal; a first channel defined by a first end of said first iimer seal and a first end of said second inner seal; a second chaimel defined by a second end of said first inner seal and a second end of said second inner seal; first and second outlet ports in said lower sheet and in fluid communication with said first and second channels, respectively, for removing a select one of said components; an mlet tabe in fluid communication with said let port, wherein said inlet tabe is fixed to the outer surface of said lower sheet and extends from the center ofthe core of said centrifuge bag; and an outlet tube having first and second ends in fluid communication with said first and second outlet ports, respectively, wherein said outlet tabe is fixed to the outer surface of said lower sheet and extends across the diameter of said core.

88. The cenfrifiige bag of claim 87, wherein said outer perimeter together with said first and second inner seals define an outer compartment.

89. The centrifuge bag of claim 87, wherein said inner perimeter together with said first and second inner seals define an inner compartment.

90. The centrifuge bag of claim 87, wherein said tubes are fluidly connected to a multiple lumen tubing comprising at least one inlet lumen and at least one outlet lumen.

91. The cenfrifuge bag of claim 90, wherein said inlet tabe is fluidly connected at its proximal end to said inlet lumen.

92. The cenfrifuge bag of claim 90, wherein said outlet tabe is fluidly connected at its axis to said outlet lumen.

93. The centrifuge bag of claim 87, wherein said fluid sample is anticoagulated whole blood.

94. The centrifuge bag of claim 93, wherein said whole blood separates radially into an outer red blood cell fraction and an inner platelet rich plasma fraction.

95. A flexible centrifuge bag for separating components of a fluid sample, comprising: upper and lower flexible sheets, said sheets having doughnut shaped configurations, imier perimeters defining a central core of said centrifuge bag and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer perimeters thereby defining the outer perimeter of said cenfrifiige bag; a first C-shaped iimer seal between said outer perimeter and said inner perimeter; a second C-shaped imier seal between said outer perimeter and said inner perimeter, wherem the concave side of said first imier seal faces the concave side of said second inner seal; a first chamiel defined by a first end of said first inner seal and a first end of said second imier seal; a second channel defined by a second end of said first inner seal and a second end of said second inner seal; an inlet tabe for introducing said fluid into said first centrifuge bag, wherein said inlet tube is sealed between said upper and lower sheets and extends from the center ofthe core of said centrifuge bag; and an outlet tube having first and second ends in fluid communication with said first and second channels, respectively, wherein said outlet tabe is sealed between said upper and lower sheets and extends across the diameter of said core.

96. The cenfrifiige bag of claim 95, wherein said outer perimeter together with said first and second inner seals define an outer compartment.

97. The cenfrifiige bag of claim 95, wherein said inner perimeter together with said first and second inner seals define an inner compartment.

98. The centrifuge bag of claim 95, wherein said tabes are fluidly connected to a multiple lumen tubing comprising an inlet lumen and an outlet lumen.

99. The centrifuge bag of claim 98, wherein said inlet tabe is fluidly connected at its proximal end to said inlet lumen.

100. The cenfrifuge bag of claim 98, wherein said outlet tabe is fluidly coimected at its axis to said outlet lumen.

101. The centrifuge bag of claim 95, wherein said fluid sample is anticoagulated whole blood.

102. The cenfrifuge bag of claim 1 , wherein said centriftige bag comprises concentric indicator lines imprinted on said centrifuge bag for monitoring the separation of said whole blood components, and wherein the distal end of said outlet tabe is positioned relative to a predetemiined indicator line.

103. The centrifuge bag of claim 30, wherein said cenfrifuge bag comprises concentric indicator lines imprinted on said centrifuge, bag for monitoring the separation of said whole blood components, and wherem the distal end of said outlet tabe is positioned relative to a predetermined indicator line.

104. The cenfrifiige bag of claim 38, wherein said centrifuge bag comprises concentric indicator lines imprinted on said cenfrifuge bag for monitoring the separation of said whole blood components, and wherem the distal end of said outlet tabe is positioned relative to a predetermined indicator line.

105. The cenfrifiige bag of claim 47, wherein said centrifuge bag comprises concenfric indicator lines imprinted on said centriftige bag for monitoring the separation of said whole blood components, and wherein the distal end of said outlet tube is positioned relative to a predetemiined indicator line.

106. The cenfrifuge bag of claim 55, wherein said cenfrifuge bag comprises concentric indicator lines imprinted on said centrifuge bag for monitoring the separation of said whole blood components, and wherein the distal end of said outlet tabe is positioned relative to a predetermined indicator line.

107. The centrifuge bag of claim 82, wherein said centrifuge bag comprises concentric indicator lines imprinted on said centrifuge bag for monitoring the separation of said whole blood components, and wherein the distal end of said outlet tabe is positioned relative to a predetemiined indicator line.

108. The centrifuge bag of claim 87, wherein said centrifuge bag comprises concentric indicator lines imprinted on said centriftige bag for monitoring the separation of said whole blood components, and wherein the distal end of said outlet tabe is positioned relative to a predetemiined indicator line.

109. The centrifuge bag of claim 95, wherein said cenfrifiige bag comprises concentric indicator lines imprinted on said centrifuge bag for monitoring the separation of said whole blood components, and wherein the distal end of said outlet tabe is positioned relative to a predetermined indicator line.

110. A method of harvesting cellular components from anticoagulated whole blood, said method comprising: a) providing a flexible centrifuge bag comprising: i) at least one inlet tabe; ii) at least one outlet tabe; iii) upper and lower flexible sheets, said sheets having doughnut shaped configurations, iimer perimeters defining a cenfral core of said centrifuge bag and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer perimeters thereby defining the outer perimeter of said centrifuge bag, and wherein said tubes are sandwiched between said upper and lower sheets and extend from the central core of said centrifuge bag, whereby when said upper and lower sheets are sealed together at their imier perimeters said tabes are sealed between said upper and lower sheets at the imier perimeter; b) providing a centrifuge rotor; c) securing said flexible centrifuge bag horizontally in said centrifuge rotor whereby said cenfrifiige bag is rotatable therewith; d) rotating said centrifuge rotor; e) providing an aliquot of said whole blood to said rotating centrifuge bag via said inlet tabe, whereby said whole blood separates radially into an outeπuost first fraction containing red blood cells, a second fraction containing white blood cells, a third fraction containing platelet rich plasma, and an innermost fourth fraction containing platelet poor plasma, wherein the distal end of said outlet tabe is in flow communication with said desired cellular component; and f) withdrawing said cellular component fraction from said rotating cenfrifuge bag via said outlet tube until a substantial portion of said cellular component fraction has been removed from said bag; and g) terminating withdrawal of said cellular component when said undesired cellular component fraction is detected to be near the distal end of said outlet tube.

111. The method of claim 110, wherein said inlet tube has a distal end positioned at a first radial distance from the center axis of said centrifuge bag and an outlet tabe having a distal end positioned at a second radial distance from the center axis of said cenfrifuge bag, said first radial distance being greater than said second radial distance.

112. The method of claim 110, wherein said cellular component fraction is withdrawn from said bag by applying suction to said outlet tabe.

113. The method of claim 110, further comprising: h) disconnecting said inlet tube from said whole blood source; i) withdrawing the remaining components from said bag by applying suction to said inlet tube; and j) repeating steps e) tlirough i) until a desired quantity of said cellular component fraction has been collected.

114. The method of claim 110, wherein said tabes are fluidly connected to a multiple lumen tubing comprising at one inlet lumen and at least one outlet lumen.

115. The method of claim 114, wherein said inlet tube is fluidly connected at its proximal end to said inlet lumen.

116. The method of claim 114, wherein said outlet tabe is fluidly connected at its proximal end to said outlet lumen.

117. The method of claim 114, wherein said multiple lumen tubing is prevented from completely rotating about its axis during rotation of said centriftige rotor.

118. The method of claim 110, wherein said detection is accomplished visually or by a sensor.

119. The method of claim 110, wherein said centrifuge rotor comprises a fransparent cover comprising one or more spaced apart concentric indicator lines for monitoring the separation of said whole blood components, and wherein the distal end of said outlet tube is positioned relative to a predetemiined indicator line.

120. The method of claim 110, wherein said cover comprises indicator lines having radial distances that indicate the location ofthe desired cellular components with respect to the distal end of said outlet port.

121. The method of claim 120, wherein the distal end of said outlet tube is positioned relative to said second indicator line.

122. A method of collecting cellular components from whole blood, said method comprising: a) providing a flexible centrifuge bag comprising at least one inlet tube, a first outlet tabe, a second outlet tabe, and a third outlet tabe; b) providing a centrifuge; c) securing said flexible cenfrifiige bag in said cenfrifiige whereby said centrifuge bag is rotatable therewith; d) rotating said centriftige and said centriftige bag; e) providing an aliquot of said whole blood to said rotating centrifuge bag via said inlet tabe, whereby said whole blood components separate radially into an outermost first fiaction containing red blood cells, a second fraction containing white blood cells, a third fraction containing platelet rich plasma, and an innermost fourth fraction containing platelet poor plasma, wherein the distal end of said outlet tube is in flow communication with said desired cellular fraction; and f) withdrawing said desired cellular fraction from said rotating centrifuge bag via said first outlet tube until a substantial portion of said desired cellular fraction has been removed from said bag and terminating withdrawal of said desired cellular fraction when said undesired cellular fraction is detected to be near the distal end of said first outlet tabe.

123. The method of claim 122, further comprising: g) repeating steps d) through f) until the volume ofthe second fraction and/or the first fraction accumulates to the point where the desired cellular fraction within the centrifuge bag is no longer in flow communication with the distal end of said first outlet tube; h) shifting the location ofthe desired cellular fraction within the cenfrifiige bag until the desired cellular fraction is in flow communication with the distal end of said first outlet tabe; and i) repeating steps d)-h) until the desired quantity of desired cellular fraction is collected.

124. The method of claim 123, wherein the distal end of said second outlet tabe is near the outer perimeter of said cenfrifiige bag, and said shifting comprises withdrawing said first fraction from said rotating centrifuge bag through said second outlet tube.

125. The method of claim 122, wherein said tabes are connected at their proximal ends to a multiple lumen tubing.

126. The method of claim 125, wherein said multiple lumen tubing comprises an inlet lumen connected at one end to said inlet tabe and at the other end to a whole blood source.

127. The method of claim 125, wherein said multiple lumen tubing comprises a first outlet lumen connected at one end to said first outlet tube and at the other end to a receiving container.

128. The method of claim 125, wherein said multiple lumen tubing is prevented from completely rotating about its axis during rotation of said cenfrifuge.

129. The method of claim 122, wherein said detection is accomplished visually or by a sensor.

130. A method for harvesting platelets from whole blood, comprising: a) providing a disposable centrifuge bag comprising: i) upper and lower flexible sheets, said sheets having doughnut shaped configurations, imier perimeters defining a cenfral core of said centrifuge bag, and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer and inner perimeters; ii) a first processing compartment for receiving said whole blood and separating said whole blood into a red blood cell fraction and a platelet suspension fraction; iii) a second processing compartment for separating platelets from said platelet suspension fraction; iv) channels for directing said platelet suspension fraction from said first processing compartment into said second processing compartment; v) an inlet tabe for infroducing said whole blood to said first processing chamber; and vi) an outlet tabe for removing platelets from said second processing compartment; b) providing a centrifuge; c) securing said flexible centrifuge bag in said cenfrifiige, whereby said cenfrifuge bag is fixably rotatable with said cenfrifiige; d) rotating said centrifuge; e) providing an aliquot of whole blood to said rotating cenfrifiige bag via said inlet tube, whereby the whole blood components separate radially within said fust processing compartment into a red blood cell fraction and a platelet suspension fraction; f) repeating step e) as necessary until said platelet suspension flows through said channels into said second processing compartment and terminating said providing step when said red blood cell fraction is detected to be near said channels, whereby said platelets separate from said platelet suspension in said second processing compartment; and g) removing said platelet concentrate from said second processing compartment via second outlet tabe.

131. The method of claim 130, wherein said inlet tabe extends radially from said cenfral core and is sealed between die upper and lower sheets at the inner perimeter.

132. The method of claim 130, wherein said second processing compartment further comprises first and second diametrically opposed wells on the inner surface for collecting said platelets.

133. The method of claim 132, wherein said outlet tube extends across the diameter of said core, wherein the first and second ends of said outlet tabe are proximal to said first and second wells, respectively.

134. The method of claim 131, wherein the first and second ends of said outlet tabe are proximal to said first and second indentations, respectively.

135. The method of claim 130, wherein said inlet tabe is fluidly comiected at its proximal end to an inlet lumen of a multi lumen tubing.

136. The method of claim 130, wherein said outlet tube is fluidly comiected at its axis to an outlet lumen of a dual lumen tubing,

137. The method of claim 135, wherem said inlet lumen is fiirther comiected to a whole blood source.

138. The method of claim 137, further comprising: h) disconnecting said inlet lumen from said whole blood source; i) withdrawing the remaining components from said bag by applying suction to said inlet lumen; j) reconnecting said inlet lumen to said whole blood source; and k) repeating steps e) tlirough j) until a desired amount of said platelets has been harvested.

139. The method of claim 130, wherein said detection is accomplished visually or by a sensor.

140. A method for harvesting platelet rich plasma from whole blood, comprising: a) providing a disposable cenfrifuge bag comprismg: i) upper and lower flexible sheets, said sheets having doughnut shaped configurations, imier perimeters defining a cenfral core of said centrifuge bag, and outer perimeters, wherein said upper and lower sheets are superimposed and completely sealed together at their outer and inner perimeters; ii) an outer compartment for receiving said whole blood; iii) an inner compartment between said outer compartment and said core; iv) first and second channels for providing fluid communication between said outer and said compartments; v) an inlet port in said lower sheet in fluid communication with said outer compartment for introducing whole blood into said outer compartment; vi) first and second outlet ports in said lower sheet and positioned in said first and second channels, respectively, for removing platelets from said cenfrifiige bag; vii) an inlet tube fixed to the outer surface of said lower sheet and extending radially from the cenfral core, wherein the distal end of said inlet tube is in fluid communication with said inlet port; and viii) an outlet tabe fixed to the outer surface of said lower sheet and extending across the diameter of said cenfral core, wherein the first and second ends of said outlet tube are in fluid communication with said first and second outlet ports, respectively. b) providing a centiifiige; c) securing said centrifuge bag in said centrifuge, whereby said cenfrifiige bag is fixably rotatable with said centrifuge; d) rotating said centrifuge; e) providing an aliquot of whole blood to said rotating cenfrifuge bag via said inlet tabe, whereby the whole blood components separate radially within said outer compartment into an outer red blood cell fraction and an hmer platelet rich plasma fraction; f) repeating step e) until said platelet rich plasma fraction flows into said channels; and g) withdrawing said platelet rich plasma fraction through said outlet ports until said red blood cell fiaction is detected to be near said outlet ports.

141. The method of claim 140, wherein said inlet tube is fluidly connected at its proximal end with an inlet lumen of a dual lumen tubing.

142. The method of claim 140, wherein said outlet tube is fluidly coimected at its axis with an outlet lumen of a dual lumen tubing.

143. The method of claim 141, wherein said inlet lumen is further connected to an anticoagulated whole blood source.

144. The method of claim 143, further comprising: h) disconnecting said mlet lumen from said whole blood source; i) withdrawing the remaining components from said bag by applying suction to said inlet lumen; j) reconnecting said inlet lumen to said whole blood source; and k) repeating steps e) through j) until a desired amount of said platelets has been harvested.

145. The method of claim 140, wherein said detection is accomplished visually or by a sensor.

146. A method of harvesting platelet rich plasma from anticoagulated whole blood, said method comprising: a) providing a rigid cenfrifiige container comprising i) a rigid annular body having an axial core; ii) an interior collection chamber for receiving and holding said whole blood during cenhifugation, said chamber having an outer perimeter and an inner perimeter and having a generally off-centered figure eight-shaped side cross-sectional configuration wherein the nanowest poition of said chamber is closer to the inner perimeter; iii) an inlet channel extending radially from said core to a distance near the outer perimeter of said chamber, the distal end of said inlet chaimel being in fluid communication with the interior of said chamber; and iv) an outlet channel extending radially from said core to a distance approximately near the nanowest portion of said chamber, said outlet channel being in fluid communication with the interior of said chamber at the nanowest portion; b) providing a cenfrifiige; c) securing said rigid centrifuge container to said cenfrifiige; d) rotating said cenfrifuge and said cenfrifuge container; e) providing an aliquot of said whole blood to said rotating cenfrifiige container via said inlet channel, whereby said whole blood components separate radially within said chamber into an outermost first fraction containing red blood cells, a second fraction containing white blood cells, a third fraction containing platelet rich plasma, and an innermost fourth fraction containing platelet poor plasma, and wherein the distal end of said outlet chaimel is positioned to be in flow conmiunication with said platelet rich plasma fraction; and f) removing said platelet rich plasma from said rotating centrifuge container via said outlet channel.

147. The method of claim 146, wherein said inlet channel and said outlet channel are fluidly coimected at their proximal ends to a dual lumen tubing having an inlet lumen and an outlet lumen.

148. The method of claim 147, wherein said inlet lumen is fluidly coimected at one end to said inlet channel and at the other end to a whole blood source.

149. The method of claim 148, further comprising: g) disconnecting said inlet lumen from said whole blood source; h) withdrawing the remaining fractions from said chamber by applying suction to said inlet lumen; i) repeating steps e)-h) as needed until the desired amount of said platelet rich plasma has been harvested.

150. The method of claim 147, wherein said dual lumen tubing is prevented from completely rotating about its axis during rotation of said centrifuge.

151. A method for harvesting platelet rich plasma from anticoagulated whole blood, comprising: a) providing a flexible cenfrifuge bag comprising an inlet tube and at least one outlet tube; b) providing a rotatable rotor comprising an annular base having a lower aimular channel and an annular cover having an upper aimular channel for holding said flexible centrifuge bag therebetween; c) positioning said flexible centrifuge bag in said base; d) superimposing said cover over said base, whereby said lower and upper lower channels define an interior chamber having a generally off-centered figure eight cross-sectional configuration and wherein said centrifuge bag is fixably rotatable with said rotor; e) rotating said rotor; f) introducing an aliquot of said fluid to said rotating centriftige bag via said inlet tube, whereby whole blood components separate into at least an outer red blood cell fraction and an inner platelet rich plasma fraction based on their densities at vaiying radial distances from the cenfral axis; and g) removing said platelet rich plasma fraction via said outlet tube.

152. The method of claim 151, wherein said base comprises a first outer rim extending from the base top surface and a first column axially centered and extending from the base top surface, wherein said first colunm has a first groove extending across the diameter of its top surface, wherein said lower aimular channel is defined by said first rim and said first colunm.

153. The method of claim 152, wherein said base column extends through said bag cenfral core.

154. The method of claim 151, wherein said cover comprises a second outer rim extending from the cover bottom surface and a second column axially centered and extending from the cover bottom surface, wherein said second column has a second groove extending across the diameter of its top surface, wherein said upper annular channel is defined by said second rim and said second colunm.

155. The method of claim 151, wherein said cenfrifuge bag further comprises superimposed upper and lower flexible sheets having doughnut shaped configurations, inner perimeters defining a cenfral core of said centrifuge bag, and outer perimeters, wherein said upper and lower sheets are completely sealed together at their outer and inner perimeters.

156. The method of claim 151, wherein said iimer and outer tabes are sandwiched between said upper and lower sheets and sealed therebetween and extend from the cenfral core of said centrifuge bag, wherein said inlet and outlet tabes are seated in said first groove said second grooves.

157. The metiiod of claim 151, wherein said cover is fabricated from a material that is at least partially transparent.

158. The method of claim 151, further comprising one or more radially spaced apart concentric indicator lines on said cover for monitoring the separation of said whole blood components.

159. The method of claim 151, wherein said inlet and outlet tubes are comiected at their distal ends to a multiple lumen tubing.

160. The method of claim 159, wherein said multiple lumen tubing is prevented from completely rotating about a cenfral axis.

161. The method of claim 151, wherein at least one of said outlet tubes is fluidly comiected at its distal ends to a bent fitting.

162. A cenfrifiige for separating components contained in a fluid, comprising: a flexible centrifuge bag having a doughnut shaped configuration and a cenfral core; a rotatable rotor comprising an annular base having a first annular chamiel and an amiular cover having a second aimular chaimel for holding a flexible cenfrifiige bag therebetween, whereby when said base and said cover are superimposed, said first and second channels define an annular mterior chamber havmg an off-centered figure eight configuration, said cover further comprising one or more radially spaced apart concentric indicator lines for monitoring the separation of said components.

163. The cenfrifiige of claim 162, wherein said base further comprises a first outer rim extending from the base top surface and a first grooved column axially centered and extending from the base top surface, wherein the height of said first rim is equal to the height of said first column, said first rim and said first colunm defining said first annular chamiel, and wherein said cover further comprises a second outer rim extending from the cover bottom surface and a second grooved column axially centered and extending from the cover bottom surface, wherein the height of said second rim is equal to the height of said second colunm, said second rim and said second column defining said second annular chamiel.

164. The centrifuge of claim 162, wherein said flexible bag comprises superimposed upper and lower flexible sheets having doughnut shaped configurations, inner perimeters defining a cenfral core of said centiifiige bag, and outer perimeters, wherein said upper and lower sheets are completely sealed together at their outer and imier perimeters, said centrifuge bag further comprising an inlet tabe and at least one outlet tabe, said tubes sandwiched between said upper and lower sheets and sealed therebetween and extending from the central core of said cenfrifiige bag, wherein the distal end of one of said outlet tabes is positioned relative to a predetermined indicator line.

165. The cenfrifuge of claim 163, wherein when said centrifuge bag is positioned between said cover and base and wherein said tubes are seated in said grooves.

166. The cenfrifuge of claim 162, wherein said components separate at vaiying radial distances based on their densities.

167. The centrifuge of claim 164, wherem said fluid is whole blood and said components comprise a red blood cell fraction, a white blood cell fraction, a platelet rich plasma fraction, and a platelet poor plasma fraction.

168. The cenfrifuge of claim 167, wherein said cover comprises first, second and third indicator lines having radial distances that indicate the location ofthe components with respect to the distal end on said outlet tabe.

169. The centrifuge of claim 168, wherein the distal end of one of said outlet tubes is positioned relative to said second indicator line.

170. A method for separating components contained in a fluid, comprising: a) providing a flexible centrifuge bag comprising an inlet tabe and at least one outlet tabe; b) providing a rotatable rotor comprising an annular base and an annular cover for holding said flexible centrifuge bag therebetween, said base having a fust outer rim extending from the base top surface and a first column axially centered and extending from the base top surface, said first rim and said first column defining a lower chamber, said cover having a second outer rim extending from the cover bottom surface and a second column axially centered and extending from the cover bottom surface, said second rim and said second column defining an upper chamber, said cover further comprising one or more spaced apart concenfric indicator lines for monitoring the separation of said components, and wherein the distal end of one of said outlet tabes is positioned relative to a predetermined indicator line; c) positioning said flexible bag in said base, wherein said base colunm extends tlirough said bag cenfral core; d) superimposing said cover over said base, whereby said lower and upper chambers define an interior chamber and wherein said centrifuge bag is rotatable with said rotor; e) rotating said rotor; f) infroducing an aliquot of said fluid to said rotating centrifuge bag via said inlet tube, whereby said components separate into fractions based on their densities at varying radial distances from the central axis; and g) removing a predetemiined fraction via said outlet tube.

171. The method of claim 170, wherein said centrifuge bag comprises superimposed upper and lower flexible sheets having doughnut shaped configurations, inner perimeters defining a cenfral core of said cenfrifuge bag, and outer perimeters, wherein said upper and lower sheets are completely sealed together at their outer and inner perimeters and wherein said inlet and said outlet tubes are sandwiched between said upper and lower sheets and sealed therebetween, said tabes extending from the cenfral core of said centiifiige bag

172. The method of claim 170, wherein said first column further comprises a first groove extending the diameter of said first column, and said second colunm further comprises a second groove extending the diameter of said second column, wherein when said bag is held between said cover and base said tabes are seated in said grooves.

173. The method of claim 170, wherein said components separate at varying radial distances based on their densities.

174. The method of claim 171, wherein said fluid is whole blood and said components comprise a red blood cell fraction, a white blood cell fiaction, a platelet rich plasma fraction, and a platelet poor plasma fraction.

175. The method of claim 174, wherein said cover comprises first, second and third indicator lines having radial distances that indicate the location ofthe components with respect to the distal end on said outlet tube.

176. The method of claim 175, wherein the distal end of said outlet tube is positioned relative to said second indicator line.

177. A centrifuge for separating components contained in a fluid, comprismg: a rotatable rotor comprising an annular base having a first amiular channel and an aimular cover having a second annular chaimel for holding a flexible cenfrifuge bag therebetween, whereby when said base and said cover are superimposed, said first and second channels define an amiular interior chamber having an off-centered figure eight configuration.

178. The cenfrifuge of claim 177, wherein said base further comprises a first outer rim extending from the base top surface and a first grooved column axially centered and extending from the base top surface, wherein the height of said first rim is equal to the height of said first column, said first rim and said first column defining said first aimular chamiel, and wherein said cover further comprises a second outer rim extending from the cover bottom surface and a second grooved column axially centered and extending from the cover bottom surface, wherein the height of said second rim is equal to the height of said second colunm, said second rim and said second column defining said second annular chaimel.

179. The cenfrifuge of claim 178, wherein said flexible bag comprises superimposed upper and lower flexible sheets having doughnut shaped configurations, inner perimeters defining a central core of said centrifuge bag and outer perimeters, wherein said upper and lower sheets are completely sealed together at their outer and imier perimeters, said cenfrifiige bag further comprising an inlet tube and at least one outlet tube, said tubes sandwiched between said upper and lower sheets and sealed therebetween, said tubes extending from the cenfral core of said centrifuge bag.

180. The centrifuge of claim 179, wherein when said centrifuge bag is positioned between said cover and said base, said inlet and outlet tabes are seated in said first and second grooves.

181. The cenfrifiige of claim 177, wherein said cover is fabricated from a material that is at least partially transparent.

182. The cenfrifiige of claim 181, further comprising one or more radially spaced apart concentric indicator lines on said cover for monitoring the separation of said components.

183. The centrifuge of claim 177, wherein said fluid is whole blood.

184. The centrifuge of claim 177, further comprising a sensor assembly positioned external to the centrifuge for monitoring the separation of said components.

185. The centriftige of claim 179, wherein at least one outlet tube is fluidly comiected at its distal end to a bent fitting.

186. The cenfrifiige of claim 179, wherein said tabes are fluidly coimected at their proximal ends to a multiple lumen tubing.

187. The centrifuge of claim 186, wherein said multiple lumen tubing is prevented from completely rotating about a central axis.

188. A method for removing a predetemiined component contained in a fluid, comprising: a) providing a flexible centrifuge bag comprising a cenfral core, an inlet tube and at least one outlet tube; b) providing a rotatable rotor an aimular base having a first annular channel and an aimular cover having a second annular channel for holding a flexible cenfrifuge bag therebetween, whereby when said base and said cover are superimposed, said first and second channels define an amiular interior chamber having an off-centered figure eight configuration; c) positioning said flexible centrifuge bag in said base; d) superimposing said cover over said base, whereby said lower and upper lower chambers define an interior chamber having a generally off-centered figure eight cross-sectional configuration and wherein said cenfrifuge bag is fixably rotatable with said rotor; e) rotating said rotor; f) introducing an aliquot of said fluid to said rotating centriftige bag via said inlet tube, whereby said components separate into fractions based on their densities at vaiying radial distances from the cenfral axis; and g) removing a fraction containing said predetermined component via said outlet tube.

189. The method of claim 188, wherein said base fiirther comprises a first outer rim extending from the base top surface and a first grooved column axially centered and extending from the base top surface, wherein the height of said first rim is equal to the height of said first column, said first rim and said first column defining said first annular chamiel, and wherein said cover further comprises a second outer rim extending from the cover bottom surface and a second grooved colunm axially centered and extending from the cover bottom surface, wherein the height of said second rim is equal to the height of said second column, said second rim and said second column defining said second annular chamiel.

190. The method of claim 189, wherein said base colunm extends through said core of said centrifuge bag.

191. The method of claim 189, wherein said tabes are seated in said first and second grooves.

192. The method of claim 188, wherein said centrifuge bag comprises superimposed upper and lower flexible sheets having doughnut shaped configurations, inner perimeters defining a cenfral core of said centrifuge bag, and outer perimeters, wherein said upper and lower sheets are completely sealed together at their outer and imier perimeters.

193. The method of claim 192, wherein said inner and outer tubes are sandwiched between said upper and lower sheets and sealed therebetween and extend from the central core of said cenfrif ige bag.

194. The method of claim 188, wherein said cover is fabricated from a material that is at least partially fransparent.

195. The method of claim 194, further comprising one or more radially spaced apart concenfric indicator lines on said cover for monitoring the separation of said components.

196. The method of claim 188, wherein said fluid is whole blood.

197. The method of claim 196, wherein said predetermined component is platelet rich plasma, platelet poor plasma, white blood cells or red blood cells.

198. The method of claim 188, wherein said inlet and outlet tabes are connected at their proximal ends to a multiple lumen tubing.

199. The method of claim 198, wherem said multiple lumen tubing is prevented from completely rotating about a cenfral axis.

200. The method of claim 188, wherein at least one of said outlet tubes is fluidly connected at its distal ends to a bent fitting.