CA2309831C - Improved methods and systems for performing molecular separations - Google Patents

Improved methods and systems for performing molecular separations Download PDF

Info

Publication number
CA2309831C
CA2309831C CA002309831A CA2309831A CA2309831C CA 2309831 C CA2309831 C CA 2309831C CA 002309831 A CA002309831 A CA 002309831A CA 2309831 A CA2309831 A CA 2309831A CA 2309831 C CA2309831 C CA 2309831C
Authority
CA
Canada
Prior art keywords
polymer
acid
solution
microchannel
channel
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Expired - Fee Related
Application number
CA002309831A
Other languages
French (fr)
Other versions
CA2309831A1 (en
Inventor
Robert S. Dubrow
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Caliper Life Sciences Inc
Original Assignee
Caliper Life Sciences Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Caliper Life Sciences Inc filed Critical Caliper Life Sciences Inc
Publication of CA2309831A1 publication Critical patent/CA2309831A1/en
Application granted granted Critical
Publication of CA2309831C publication Critical patent/CA2309831C/en
Anticipated expiration legal-status Critical
Expired - Fee Related legal-status Critical Current

Links

Classifications

    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/416Systems
    • G01N27/447Systems using electrophoresis
    • G01N27/44704Details; Accessories
    • G01N27/44747Composition of gel or of carrier mixture
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/416Systems
    • G01N27/447Systems using electrophoresis
    • G01N27/44704Details; Accessories
    • G01N27/44752Controlling the zeta potential, e.g. by wall coatings
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/416Systems
    • G01N27/447Systems using electrophoresis
    • G01N27/44756Apparatus specially adapted therefor
    • G01N27/44791Microapparatus

Abstract

The present invention provides methods of electrophoretically separating macromolecular species, as well as compositions and systems useful in carrying out such methods. Specifically, the methods of the present invention comprise providing a substrate that has at least a first capillary channel disposed therein. The surface of the channel has a first surface charge associated therewith, and is filled with a water soluble surface adsorbing polymer solution that bears a net charge that is the same as the charge on the capillary surface.

Description

IMPROVED METHODS AND SYSTEMS FOR PERFORMING MOLECULAR
SEPARATIONS

BACKGROUND OF THE INVENTION
Capillary electrophoresis has been established as a highly effective method for separating macromolecular species in order that they might be further characterized.
Protein and nucleic acid molecules are two major examples of molecular species that are routinely fractionated and characterized using capillary electrophoretic systems. These systems have generally proven effective as a result of the high surface to volume ratio of the thin capillaries. This high surface to volume ratio allows for much greater heat dissipation, which in turn, allows application of greater electrical currents to the capillary thereby resulting in a much more rapid separation of macromolecules introduced into the system.
In the capillary electrophoretic, size-based separation of biological macromolecules of interest, e.g., proteins and nucleic acids, electrophoretic separation is not possible in a free solution. Instead, such separation requires the presence of a matrix that alters the electrophoretic mobilities of these molecules based upon their relative size.
Although early capillary electrophoresis systems utilized solid gel matrices, e.g., cross-linked polyacrylamides, more recent systems have employed liquid polymer solutions as a flowable matrix, which permits adequate separation efficiencies without the drawbacks of cross-linked capillary systems, i.e., in introducing such matrices to or removing them from capillary channels.
For example, U.S. Patent No. 5,126,021 reports a capillary electrophoresis element which includes a capillary electrophoresis tube containing a low viscosity uncharged polymer solution, for separating nucleic acids.
U.S. Patent No. 5,264,101 to Demorest et ai. reports the use of a hydrophilic polymer solution, which is characterized by a molecular weight of 20 to 5,000 kD, and a charge between 0.01 and I% as measured by the molar percent of total monomer subunits to totai polymer subunits, where the charge is opposite to the charge of the surface of the capillary in which the polymer is used. This opposite charge of the polymer is reported to result in an interaction between the polymer and the capillary wall to reduce electroosmotic flow within the capillary.
U.S. Patent Nos. 5,552,028 and 5,567,292, both to Madabhushi et al., report the use of a uncharged, water soluble, silica adsorbing polymer in a capillary electrophoresis system to reduce or eliminate electroosmotic flow.
Surprisingly, the present inventor has discoveraed that polymer solutions can be used in capillary channel systems, which polymers employ a charge that is the same as that of the internal capillary surface, e.g., positive or negative. Even more surprisingly, it has been discovered that electroosmotic flow in capillary channel systems containing such polymer solutions is maintained the same level or lower than with an uncharged polymer solution. The present invention provides such polymers, as well as methods of utilizing these polymers and systems employing such polymers.

The present invention generally provides novel methods and compositions for use in the separation of molecular, and particularly macromolecular species by electrophoretic means.
For example, in an aspect of the present invention is provided a method of separating macromolecules by capillary electrophoresis. The method generally comprises providing a substrate which includes at least a first capillary channel disposed therein, where a surface of the channel has a first surface charge associated therewith. The capiilary channel is filled with a water soluble hydrophilic polymer solution which includes a percent charge of from about 0.0196 to about 2%, as calculated by the molar percent of charged monomer subunits to total monomer utilized in producing the polymer. The charged monomer subunits have a charge that is the same as the first surface charge. A
sample containing macromolecules is introducxd into one end of the capillary channel and a voltage gradient is applied across the length of the capillary channel, whereby the macromolecules in the sample are separated in the capillary channel. In preferred aspects, the surface charge of the capillary channel, as well as the charged monomer subunits bear a negative charge.
In further preferred aspects, the capillary channel is disposed within a silica substrate.
In a related aspect, the present invention also provides systems and apparatus for practicing the above methods. In particular, the present invention provides a system for separating macromolecules by capillary electrophoresis. The system comprises a substrate having at least a first walled capillary channel disposed therein, where the channel includes a net surface charge associated with its interior surfaces. A solution of silica adsorbing polymer as described above, is disposed in the capillary channel. The system also includes a power source electrically coupled to the capillary channel for applying a voltage gradient across the capillary channel.
Various embodiments of this invention provide a device, comprising: at least one microchannel, the microchannel comprising a polymer disposed therein, the polyrner comprising a net charge of between about 0.01 and 2%, the net charge being of the same charge as at least one surface of the microchannel.
Various embodiments of this invention provide a system for separating macromolecules by capillary electrophoresis, comprising: a substrate having at least a first walled capillary channel disposed therein, the channel having a net surface charge associated with interior surfaces of the channel; a solution of silica adsorbing polymer disposed in the capillary channel, the solution of polymer comprising: a molecular weight between about 1 kD and 5,000 kD; a net charge of between about 0.01 and 2%, the net charge being the same as the net surface charge; and a power source electrically coupled to the first capillary channel for applying a voltage gradient across the capillary channel.
Various embodiments of this invention provide a system for separating nucleic acids by molecular weight, comprising: a silica substrate having a walled capillary channel disposed therein, the channel having a negative charge associated with interior surfaces of the channel; a solution of silica adsorbing polymer disposed in the capillary channel, the solution of polymer comprising: a molecular weight between about 1 kD and 5,000 kD; a net negative charge of between about 0.01 and 2%; and a power source for applying a voltage gradient across the capillary channel.
Various embodiments of this invention provide a walled capillary for separating macromolecules by capillary electrophoresis, comprising: a capillary channel disposed in a solid substrate, interior surfaces of the capillary channel having a first net surface charge associated therewith; and a solution of silica adsorbing polymer disposed in the capillary channel, the polymer comprising: a molecular weight between about 1 kD and about 5,000 kD; a net charge of between about 0.01 and 2%, the net charge being the same as the first net surface charge.
Various embodiments of this invention provide a method of preparing a walled capillary channel for use in separating macromolecules, comprising:
filling the walled capillary channel with a silica adsorbing polymer solution, wherein the polymer has a net charge that is the same as a net charge associated with interior surfaces of the walled capillary channel.
Various embodiments of this invention provide a method of manufacturing a microfabricated channel system, the method comprising: providing a device comprising at least one microchannel; and, disposing a polymer in the at least one microchannel, the polymer comprising a net charge of between about 0.01 % and 2%, the net charge being of the same charge as at least one surface of the microchannel. Also provided are microfabricated channel systems prepared according to this method.
Various embodiments of this invention provide a method of separating macromolecules by capillary electrophoresis, comprising: providing a substrate comprising at least a first capillary channel disposed therein, a surface of the channel having a first surface charge associated therewith; filling said capillary channel with a water soluble hydrophilic polymer solution having a percent charge of from about 0.01 % to about 2%, as calculated by molar percent of charged monomer subunits to total monomer utilized in producing the polymer, the charged monomer subunits consist of monomer subunits having a charge that is the same as the first surface charge; introducing a sample containing the macromolecules into one end of the capillary channel and; applying a voltage gradient across the length of the capillary channel, whereby the macromolecules in the sample are separated in the capillary channel.
Various embodiments of this invention provide a method of separating macromolecules by capillary electrophoresis, comprising: providing a silica substrate having a capillary channel disposed therein, a surface of the channel having a negative surface charge associated therewith; filling said capillary channel with a water soluble hydrophilic polymer solution having a net charge of from about 0.1% to about 2%, the charge being the same as the surface charge; introducing a sample containing the macromolecules into one end of the capillary channel; and applying a voltage gradient across the length of the capillary channel, whereby the macromolecules in the sample are separated in the capillary channel.

3a BRIEF DESCRIPTION OF THE FIGURES
Figure 1 schematically illustrates a silica microscale electrophoresis device for use in electrophoretic separation of sample component for up to 12 different sample materials, in accordance with the present invention.
Figure 2 illustrates the chromatographic separation of DNA standard samples in a silica microscale integrated channel electrophoresis device first filled with a neutral polymer solution.
Figure 3 illustrates the chromatographic separation of DNA standard samples in a silica microscale integrated electrophoresis device first filled with a polymer solution having a negative charge associated with it.
Figure 4 illustrates a chromatographic separation as in Figure 3, but employing a charged polymer that has a larger average molecular weight and viscosity than the polymer solution used in generating the chromatogram shown in Figure 3.
Figure 5 illustrates a chromatographic separation as in Figure 4, except employing a polymer solution that has a still larger molecular weight and viscosity than the polymer used in generating the chromatogram shown in Figure 4.
Figure 6 illustrates a channel geometry for a planar polymeric substrate/microscale channel device used to perform macromolecular separations in accordance with the present invention.
Figure 7 illustrates a chromatographic separation of a 100bp ladder in a polymethylmethacrylate microfluidic device using a polymer of the invention.
Figures 8A and 8B show chromatograms comparing separations performed with just high molecular weight sieving polymer (Figure 8A) or a mixture of high molecular weight and low molecular weight sieving polymer (Figure 8B).

3b DETAILED DESCRIPTION OF THE INVENTION
The present invention provides methods of electrophoretically separating macromolecular species, as well as compositions and systems useful in carrying out such methods. Specifically, the methods of the present invention comprise providing a substrate that has at least a first capillary channel disposed therein. The surface of the channel has a first surface charge associated therewith, and is filled with a water soluble surface adsorbing polymer solution that bears a net charge that is similar to or the same as the charge on the capillary surface, e.g., positive or negative.
As used herein, the term substrate typically refers to a solid substrate in which a capillary channel is disposed. Exemplary substrates include silica based substrates, such as silica, e.g., glass, quartz or the like, silicon, etc., polymeric substrates, e.g., plastics like polydimethylsiloxanes (PDMS), polymethylmethacrylate (PMMA), polyurethane, polyvinylchloride (PVC), polystyrene, polysulfone, polycarbonate, polytetrafluoroethylene (TeflonTM), and a variety of others that are well known in the art. Substrates may take a variety of shapes or forms, including tubular substrates, e.g., polymer or fused silica capillaries, or the like. In preferred aspects, however, the substrate comprises a planar body structure in which grooves are fabricated to define capillary channels when overlaid with a cover element, also typically planar in structure. Examples of such planar capillary systems are described in PCT international application WO 98/49548 filed April 13, 1998.
Capillary channels also can be any of a variety of different shapes in cross-section, including tubular channels, rectangular channels, rhomboid channels, hemispherical channels or the like, or even more arbitrary shapes, such as may result from less precise fabrication teclmiques, e.g., laser ablation. Typically, the shape of a capillary channel will vary depending upon the substrate type used and the method of fabrication. For example, in typical fused silica capillaries, the capillary channel will be tubular. In systems employing planar substrates, on the other hand, channels will typically comprise either a rhomboid, rectangular or hemispherical cross sectional shape, depending upon the substrate material and method of fabrication of the channels.
A variety of manufacturing techniques are well known in the art for producing microfabricated channel systems. For example, where such devices utilize substrates commonly found in the semiconductor industry, manufacturing methods regularly employed in those industries are readily applicable, e.g., photolithography, wet chemical etching, chemical vapor deposition, sputtering, electroforming, etc.
Similarly, methods of fabricating such devices in polymeric substrates are also readily available, including injection molding, embossing, laser ablation, LIGA techniques and the like.
Other useful fabrication techniques include lamination or layering techniques, used to provide intermediate microscale structures to define elements of a particular microscale device.

Typically, the capillary channels will have an internal cross-sectional dimension, e.g., width, depth, or diameter, of between about 1 m and about 500 m, with most such channels having a cross-sectional dimension in the range of from about 10 m to about 200 m.

In particularly preferred aspects, planar microfabricated devices employing multiple integrated microscale capillary channels are used. Briefly, these planar microscale devices employ an integrated channel network fabricated into the surface of a planar substrate. A second substrate is overlaid on the surface of the first to cover and seal the channels, and thereby define the capillary channels.
One or more analysis channels are provided in the device with additional channels connecting the analysis channel to multiple different sample reservoirs. These reservoirs are generally defined by apertures disposed in the second overlaying substrate, and positioned such that they are in fluid communication with the channels of the device. A
variety of specific channel geometries are employed to optimize channel layout in terms of material transport time, channel lengths and substrate use. Examples of such microscale channel network systems are described in PCT international application WO
98/49548 filed April 13, 1998. One specific example of a channel geometry is illustrated in Figure 1. In operation, sample materials are placed into one or more of the sample reservoirs 116-138.
A first sample material, e.g., disposed in reservoir 116, is then loaded by electrokinetically transporting it through channels 140 and 112, and across the intersection with the separation channel 104, toward load/waste reservoir 186 through channel 184.
Sample is then injected by directing electrokinetic flow from buffer reservoir 106 through analysis channel 104 to waste reservoir 108, while pulling back the sample in the loading channels 112:114 at the intersection. While the first sample is being separated in analysis channel 104, a second sample, e.g., that disposed in reservoir 118, is preloaded by electrokinetically transporting it into channels 142 and 112 and toward the load/waste reservoir 184 through channel 182. After separation of the first sample, the second sample is then loaded across the intersection with analysis channel 104 by transporting the material toward load/waste reservoir 186 through channel 184.
The interior surface of the capillary channels typically has a charge associated with it. For example, in the case of capillary channels disposed in silica-based substrates, e.g., glass or quartz, the interior surface of the channel typically includes negatively charged chemical groups, e.g., silane groups, associated with it.
Similarly, polymeric substrates also typically comprise some level of charged chemical groups at their surface, although at much lower level than in the case of silica-based substrates. As used herein, a "charged surface" of a capillary is typically characterized by its ability to support an electroosmotic mobility of a fluid or material in the channel. In particular, channels having charged surfaces as described herein, are typically capable of supporting an electroosmotic mobility ( EO) of at least about 1 X 10-5cm2V-ls 1, for a buffer when that buffer is in contact with those walls, e.g., disposed within those channels, e.g., a buffer of from about 1 mM to about 100 mM sodium borate at a pH of from about 6 to about 9. For the purposes of the present invention, EO is defined in terms of a standard buffer of from about 1 mM to about 10 mM sodium borate buffer, at a pH of from about 7 to about 9, for example, 5 mM sodium borate, pH 7. In more common aspects, the charged surfaces in contact with the fluid are capable of supporting a pEO under the above conditions, of at least about 2 X 10"5cm2V-ts"1, preferably, at least about 5 X 10-5cm2V-1 s 1, and in particularly preferred aspects, at least about 1 X 10"6 cmzV's 1.
Different surfaces can also be treated to present differing levels or types of charged groups. Examples of such surface treatments are described in detail in PCT
international application WO 98/46438 filed April 14, 1998. In particularly preferred aspects of the present invention, capillary channels disposed in silica substrates are used, e.g., planar silica substrates or fused silica capillaries.
In aqueous systems, when charged capillary surfaces are combined with electric fields necessary for electrophoretic separation, electroosmotic flow results. For many separations, e.g., protein separations, some electroosmotic flow is actually desired, in order to ensure a net movement of all proteins through a capillary channel and past a detector. However, it is generally desirable to be able to precisely control that level of flow.
In the capillary electrophoretic separation of nucleic acids on the other hand, it is generally desirable to suppress electroosmotic flow entirely, to enhance resolution of separation.
Further, such charged surfaces have been implicated in the binding of components of samples, e.g., proteins, etc., which binding has been blamed for reduced efficiency of separation.
In accordance with the methods of the present invention, the above described capillary channel or channels are filled with a solution of a water-soluble silica-adsorbing polymer. The polymer typically includes a percent charge of between about 0.01% and 2%
that is the same as the charge that is associated with the interior wall surface of the capillary channel. By "a charge that is the same as the charge of the interior surface of the capillary channel" is meant that the polymer includes charged monomer subunits that are the same charge, e.g., negative or positive, as the charged chemical groups on the interior surface of the capillary channel. Thus, where a capillary channel includes negatively charged groups on the interior surface, e.g., silane groups in silica capillary channels, the polymer will include monomer subunits that are negatively charged. In accordance with the present invention, the polymer will preferably not include any charged monomer subunits that have a charge opposite to the charge on the interior surface of the capillary channel. In preferred aspects, the polymer has a percent charge of between about 0.01% and about 1%, more preferably, between about 0.01 % and about 0.5 %, and still more preferably between about 0.05 % and 0.5 %, and often between about 0.05% and 0.2%. As noted above, in preferred aspects, the present invention utilizes silica based substrates, e.g., planar substrates or capillaries. As such, also in preferred aspects, the polymers used in accordance with the invention are negatively charged, as is the interior surface of the capillary channel.
As used herein, the "percent charge" of a polymer refers to the molar percent of charged monomer units to total monomer subunits used in the synthesis of the polymer.
Thus, if the synthesis reaction is carried out by mixing 1 mmol of charged subunit and 99 mmol of uncharged monomer subunit, the polymer would have a percent charge of 1%, as defined herein.
The water soluble polymers of the present invention are preferably surface adsorbing polymers, and more preferably, silica adsorbing polymers, e.g., as that term is defined in U.S. Patent No. 5,567,292. Examples of particularly preferred surface adsorbing polymers include acrylic polymers, e.g., polyacrylamides, polymethylacrylamides, polydimethylacrylamides, and the like. Each of these polymers is readily synthesized to incorporate charged monomer subunits bearing a charge that is the same as the charge of the interior surface of the capillaries, e.g., negatively charged subunits. For example, carboxylic acid monomers can be used to impart a negative charge to the polymer. Such monomers include, e.g., acrylic acid, bisacrylamidoacetic acid, 4, 4-Bis(4-hydroxyphenyl)pentanoic acid, 3-butene-1, 2, 3-tricarboxylic acid, 2-carboxyethylacrylate, itaconic acid, methacrylic acid, 4-vinylbenzoic acid, and others. Sulfonic acid or phosphoric acid monomers may also be used to impart negative charge, including, e.g., 2-acrylamido-2-methyl-l-propanesulfonic acid, 2-methyl-2-propene-l-sulfonic acid, 2-propene-l-sulfonic acid, 4-styrenesulfonic acid, 2-sulfoethyl methacrylate, 3-sulfopropyldimethyl-3-methacrylamidopropylammonium inner salt, 3-sulfopropyl methacrylate, vinylsulfonic acid, Bis(2-methacryloxyethyl)phosphate, monoacryloxyethyl phosphate, and others. In the case of systems employing capillary channels with positively charged surfaces, positively charged monomer units are substituted. A variety of such subunits are known to those of skill in the art, and include, for example, quaternary amine monomers, such as 2-acryloxyethyltrimethylammonium chloride, diallyldimethylammonium chloride, 2-methacryloxyethyltrimethylammonium chloride, 3-methacryloxy-2-hydroxypropyltrimethylammonium chloride, and others.
In particularly preferred aspects, the surface adsorbing polymer is a polydimethylacrylamide polymer-co-acrylic acid. In this case, the polymer is a dimethylacrylamide polymer incorporating a desired percentage of charged acrylic acid monomers, as described above.
Synthesis of polymers used in the methods of the present invention may be carried out by any number of methods that are well known in the art.
Typically, synthesis conditions and protocols will vary depending upon the polymer to be synthesized and the nature and amount of charge to be incorporated. Examples of suitable polymer synthesis methods are described in, e.g., Odian, Principles of Polymerization, Third Ed.
(John Wiley, new York, 1991), and U.S. Patent Nos. 5,264,101 and 5,567,292.
For use, the polymer may be provided in an aqueous solution at a concentration between about 0.01 % and 30% (w/v). Different concentrations may be used depending upon the nature of the separation to be performed, the size of the capillary channel and the like. Preferably, the polymer concentration, as used in the separation methods described herein, is between about 0.0 1% and about 20% (w/v), and more preferably, between about 0.1 % and about 10%.
The average molecular weight of the polymer within the polymer solutions may vary somewhat depending upon the application for which the polymer solution is desired. For example, applications which require higher resolution, e.g., single base resolution in sequencing applications, may utilize higher molecular weight polymer solutions, while less stringent applications can utilize lower molecular weight polymer solutions. Typically, the polymer solutions used in accordance with the present invention have an average molecular weight in the range of from about 1 kD to about 6,000 kD, "preferably between about 1 kD and about 1000 kD, and more preferably, between about 100 kD and about 1000 kD.
Additionally, depending upon the particular application for which the polymer solution is being used, one may use a combination of different molecular weights, in order to capitalize on the benefits of each type of polymer. For example, higher molecular weight polymers typically provide better resolution for larger moleailes, while lower molecular weight polymer solutions provide better resolution for smaller molecules.
As such, for broader spectrum separations, i.e., having very large and very small molecules to be separated, it is often useful to incorporate both lower and higher molecular weight polymers in the overall separation solution that is being used. Typically, the ratio of high molecular weight polymer to low molecular weight polymer varies depending upon the material to be separated. Generally, however, the ratios of high molecular weight polymer to lower molecular weight polymer range from about 1:10-to about 10:1, and preferably, from about 5:1 to about 1:5 and more preferably, from about 1:2 to about 2:1. Typically, the high molecular weight polymer component has an average molecular weight of between about 300 kD and about 1,000 kD, and preferably, between about 400 kD and about 600 kD. The low molecular weight polymer, on the other hand, typically has an average molecular weight of between about 50 kD and about 300 kD, and preferably, between about 50 kD and about 200 kD. The overall percentage of polymer in the separation solution still remains in the ranges described herein.
In addition to the percent charge and molecular weights described above, the polymers used in accordance with the present invention are also characterized by their viscosity. In particular, the polymer components of the system described herein typically have a solution viscosity as used within the capillary channel, in the range of from about 2 to about 1000 centipoise, preferably, from about 5 to about 200 centipoise and more preferably, from about 10 to about 100 centipoise.
In addition to the polymer component, the polymer solution typically includes buffers for controlling pH and conductivity, other polymers and the like, as necessary for accomplishing the desired separation, i.e., neutral polymers for enhancing sieving, and the like.
In operation, a solution of the water-soluble surface-adsorbing polymer is introduced into the capillary channel. This introduction may be as simple as placing one end of the channel into contact with the polymer solution and allowing the polymer to wick into the channel. Alternatively, vacuum or pressure may be used to drive the polymer solution into the capillary channel. In the preferred integrated channel systems, the polymer solution is typically placed into contact with a terminus of a common capillary channel, e.g., a reservoir disposed at the end of a separation channel, and slight pressure is applied to force the polyrner into all of the integrated channels.
The sample containing the macromolecular species for which separation is desired, is placed in one end of the separation channel and a voltage gradient is applied along the length of the channel. As the sample components are electrokinetically transported down the length of the channel and through the polymer solution disposed therein, those components are resolved. The separated components are then detected at a point along the length of the channel, typically near the terminus of the separation channel distal to the point at which the sample was introduced.
Detection of separated species is typically carried out using UV, amperometric and/or fluorescent detection systems that are well known in the art.
Typically, such detection systems operate by detecting a characteristic optical property of the macromolecular species of interest, e.g., UV absorbance of double bonded structures, fluorescence of an associated labeling moiety, light scattering, etc. For example, in the case of fluorescent detection, such detection systems typically employ a fluorescent or fluorogenic-labeling group coupled to the various macromolecules. For instance, in the case of nucleic acids, a variety of fluorescent labeling techniques can be used. These are generally well known in the art, and include the use of covalently attached fluorescent labeling groups, e.g., as described in U.S. Patent Nos. 4,711,955, 5,171,534, 5,187,085, 5,188,934, and 5,366,860. Alternatively, associative labeling groups may be used, which preferentially associate with the macromolecular species of interest, or are only detectable, e.g., fluorescent or fluorogenic, when associated with the macromolecules of interest.
Examples of such labeling groups include, e.g., intercalating dyes for double stranded nucleic acids, streptavidin/biotin labeling groups.
As noted, preferred aspects of the present invention utilize fluorescent detection systems. Typically, such systems utilize a light source capable of directing light energy at the separation channel as the separated macromolecular species are transported past. The light source typically produces light of an appropriate wavelength to activate the labeling group. Fluoresced light from the labeling group is then collected by appropriate optics, e.g., an objective lens, located adjacent the capillary channel, and the collected light is directed at a photometric detector, such as a photodiode or photomultiplier tube. The detector is typically coupled to a computer, which receives the data from the detector and records that data for subsequent storage and analysis.
The polymer compositions are widely applicable in the separation of macromolecular species using electrophoretic techniques. Such macromolecular species include without limitation, nucleic acids, proteins, peptides, carbohydrates, and the like. In particularly preferred aspects, the polymer compositions described herein are used in the electrophoretic separation and/or identification of nucleic acids in a sample.
Such nucleic acids may include fragments or portions of genomic DNA, e.g., for genotyping, fragments or portions of mRNA, e.g., for gene expression analysis, or polymerization reaction products for verification of amplification processes. In addition, such polymer compositions are particularly useful in separating nested sets of nucleic acid fragments or synthesis products, for determination of nucleotide sequence, e.g., as prepared in Sanger or Maxam and Gilbert sequencing operations. In these sequencing operations, the nested sets of fragments typically include a number of fragments of a target nucleic acid sequence that differ in length from the next fragment by a single nucleotide, e.g., a single base extension.
The fragments in these nested sets are then separated by size in, e.g., capillary electrophoretic operations, and characterized by their terminal nucleotides.
Analysis of all of the nested fragments then provides the nucleotide sequence of the target sequence.
Examples of preferred sequencing operations are described in, e.g., U.S.
Patent 5,171,534, which employ four differentially labeled dideoxynucleotides in a Sanger sequencing operation. Each labeled dideoxynucleotide has a different fluorescent emission or absorption maximum. Random incorporation of each of the four dideoxynucleotides during target template dependent polymerization results in a nested set of fragments including all possible extension products, where each extension product is differentially labeled by virtue of its terminal dideoxynuclotide. The differential labeling permits characterization of the terminal nucleotide in a single detection operation, and subsequent determination of the overall sequence of the target nucleic acid.
The present invention is further illustrated by the following non-limiting examples.

WO 99/31495 PCTNS98lZ5362 EXAMPLES
I. Po vmer Synthesis Polymer solutions were prepared according to the following protocols:
A. 2-Methyl-I -p,=anql pQl rization of p,W,XdiMr"acrvlamide To a 25 ml sidearm flask was added 6 ml of 2-mcthyl-l-propanol and 3 ml of N,N-dimethylacrylamide. The flask was fitted with a one-hole rubber stopper that had an argon gas line feed through the hole to the bottom of the flask. The side arm of the flask was left open. A steady stream of argon was bubbled through the solution in the flask for minutes. After the 10 minute bubbling period was over three milligrams of 10 2,2'Azobisisobutyronitrile was added to the flask. The flask was lowered into a 60 C water bath and the bubbling of the argon gas continued. After one hour the flask was allowed to cool to ambient temperature. The solution in the flask was now a viscous liquid indicating that polymerization had occurred. Purification of the polynzer was achieved by subjecting it to a series of precipitation's and dissolutions. The polymer was precipitated out in 100 nil of hexane. The hexane was poured off and the polymer precipitate was dissolved in about 50 ml of inethylene chloride. This solution was then precipitated out in hexane again and redesolved in methylene chloride. After one final hexane precipitation the purified polymer was vacuum dried for 48 hours. It was then stored in a glass vial labeled Polymer #1.
B. 2-Methyl-l-pDpanoJ pQlyMerization of pQl,ydimethvlacrvl~rylic acid ( .9/0.1) To a 25 ml sidearm flask was added 6 ml of 2-methyl-l-propanol, 3 ml of N,N-dimethylacrylamide and 0.0054 ml of acrylic acid. The flask was fitted with a one-hole rubber stopper that had an argon gas line feed through the hole to the bottom of the flask.
The side arm of the flask was left open. A steady stream of argon was bubbled through the solution in the flask for 10 minutes. After the 10 minute bubbling period was over three milligrams of 2,2'Azobisisobutyronitrile was added to the flask. The flask was lowered into a 60 C water bath and the bubbling of the argon gas continued. After one hour the flask was allowed to cool to ambient temperature. The solution in the flask was now a viscous liquid indicating that polymerization had occurred. Purification of the polymer was achieved by subjecting it to a series of precipitation's and dissolutions. The polymer was precipitated out in 100 ml of hexane. The hexane was poured off and the polymer precipitate was dissolved in about 50 ml of methylene chloride. This solution was then precipitated out in hexane again and redesolved in methylene chloride. After one final WO 99/31495 PCTNS98f2SSG2 hexane precipitation the purified polymer was vacuum dried for 48 hours. It was then stored in a glass vial labeled Polymer #2.
C. Aau~ymerization of medium molecular weight volv 1X ac;ylamide/acrvlic acid (99.9/0.1) To a 25 ml sidearm flask was added 4.0m] of inethanol, 5.OmL of deionized water, 1.OmL of NN-dimethylacrylamide and 0.0018 ml of acrylic acid. The flask was fitted with a one-hole rubber stopper that had an argon gas line feed through the hole to the bottom of the flask. The side arm of the flask was left open. A steady stream of argon was bubbled through the solution in the flask for 10 minutes. A 10 percent solution of ammonium persulfate was made in deionized water. After the 10-minute bubbling period was over 200 l of the ammonium persulfate solution was added to the flask.
The flask was lowered into a 50 C water bath and the bubbling of the argon gas continued.
After 45 minutes the flask was allowed to cool to ambient temperature. The solution in the flask was now a viscous liquid indicating that polymerization had occurred. The solution was transferred into 10 kD dialysis tubing (Spectrum Technologies, part number 132680). The loaded tubing was placed into 1000 ml of deionized water and stirred for 24 hours. The water was replaced with fresh deionized water and stiffed for another 24 hours. After this second 24 hours was complete the dialysis bag was removed from the water, and the contents poured out into a plastic tray. The tray containing the polymer solution was placed in a 60 C oven for 4 hours to dry. The tray was then removed from the oven and the thin clear film of polydimethylacrylamide/ acrylic acid (99.9/0.1) was peeled from the tray and placed in a glass vial for storage and labeled Polymer #3.
D. Aqueous p,2lymerization of hia_h_ molecular weiQht p,glydimethylacprlamide/acrvlic acid {99 9/0 1) To a 25 mi sidearm flask was 8.0 ml of deionized water, 2.0 ml of N,N-dimethacrylamide and 0.0018 ml of acrylic acid. The flask was fitted with a one-hole mbber stopper that had an argon gas line feed through the hole to the bottom of the flask.
The side arm of the flask was left open. A steady stream of argon was bubbled through the solution in the flask for 10 minutes. A 10 % solution of ammonium persulfate was made in deionized water. After the 10-minute bubbling period was over 200 l of the ammonium persulfate solution was added to the flask. The flask was lowered into a 50 C
water bath and the bubbling of the argon gas continued. After 45 minutes the flask was allowed to cool to ambient temperature. The solution in the flask was now a soft gel-like material indicating that polymerization had occurred. The polymer was diluted with 30 ml of deionized water and then 10 ml of this solution was transferned into 10 kD
dialysis tubing (Spectrum Technologies, part number 132680). The loaded tubing was placed into 1000m1 of deionized water and stirred for 24 hours. The water was replaced with fresh deionized water and stirred for another 24 hours. After this second 24 hours was complete the dialysis bag was removed from the water, and the contents poured out into a plastic tray. The tray containing the polymer solution was placed in a 60 C oven for 4 hours to dry.
The tray was then removed from the oven and the thin clear film of polydimethacrylamide/acrylic acid (99.9/0.1) was peeled from the tray and placed in a glass vial for storage and labeled Polymer #4.
E. ViscosiIy Measurements of Polymers The viscosity of the various polymers prepared as above, was measured at C using an Ubberholde viscometer (Technical Glass Products, Dover NJ) following the ASTM D445 test method. Each polymer was mixed with water to the concentration 15 (weight/volume) at which it was used for the electrophoretic separations in the following examples. Viscosities are provided in Table I, below:

Table I

Polymer Concentration Viscosity M (Centipoise) #1 6.5 5.7 #2 6.5 7.4 #3 2.0 34.2 #4 1.8 60.1 IL Electrophoretic arations 20 The polymers, synthesized as described above, were used to perform separations of standard nucleic acid samples in a microseale integrated channel device to demonstrate their efficacy, as follows:

A. Separation of 100 bpj&ker with Control Polvmer #1 A 6.5% solution of neutral control polymer (Polynier #1) was prepared by dissolving the polymer in water at a concentration of 10% (w/v). The polymer solution for use in separations was then made up by mixing 0.65 ml of polymer solution, 0.2 ml Genetic Analysis Buffer (Perkin-Elmer, Norwalk CT), and 0.15 ml distilled water.
Intercalating dye (Syto 61, Molecular Probes, Inc.) was added to the polymer solutions at a ratio of 1:2500.
Sample buffer was prepared by adding 2 ml Genetic Analysis Buffer to 8 ml distilled water and4 lofSyto61Tm.
Experimental separations were performed on a 100 bp ladder (Promega) which contains nucleic acid fragments ranging from 100 to 1000 bp in length, at 100 bp increments, and also including a 1500 bp fragment. The samples were prepared by diluting the stock ladder solution 1:10 in the sample buffer containing Syto 61TM.
Sample separations were performed in a multi-sample microscale capillary electrophoresis device in which multiple samples are serially separated along a common separation capillary channel. The device employed a planar glass chip construction, where the channels were etched as grooves in a first planar glass substrate and a second glass .
substrate is overlaid and bonded to the first, to define the channels. The integrated channel device had the channel geometry shown in Figure 1, which allows the serial analysis of up to 12 samples along the same separation channel.
The channels of the device were filled with Polymer #1 by introducing the polymer into one common reservoir and allowing the polymer solution to wick into all of the interconnected channels. Nine sample wells in the device were filled with the sample buffer containing the ladder DNA, while three wells were filled with plain sample buffer (no DNA). The separation was run in the device using an electrical controller operating under current control. Separated species were fluoresced using a red laser diode directed at a point along the separation channel, and fluorasced light was collected by an objective lens and transmitted to a photomultiplier tube for detection. Signal was recorded on a PC as a fiwetion of retention time. The separation data obtained using the neutral polymer solution, e.g., uncharged polydimethylacrylamide polymer solution, is shown in Figure 2, as a plot of fluorescence intensity (in arbitrary units) as a function of retention time (seconds).
Specifically, Figure 2 illustrates separation of the 100 bp ladder, in three repiicate separations (Sample B 1, B3 and B4) as well as a control run in which no DNA
was introduced (Sample B2). A total of nine replicate separations and three control runs were WO 99l3149S PCr/US9sn5562 performed, and the data from each separation was virtually identical to that shown in Figure 2.
B. Sgparation of 100 bo L.adder with Poly= #2 A solution of negatively charged polymer (Polymer #2) was prepared in the same fashion as Polymer #1, in Example II A., above. This polymer solution was again used to perform a separation of an identically prepared nucleic acid sample in an identical multi-sample device under identical electrical control.
Figure 3 illustrates the data obtained using Polynn~r #2 in the identical separation (Sample B 1, B3 and B4) and control (Sample B2). Again, a total of ten replicate separations and two control runs were performed, and the data in each case was virtually identical to that shown.
C. SgpWation of a IOObo Ladder with Poly~#3 A 2.0% solution of Polymer 3 was prepared by adding 0.20g of Polymer 3, 2.Og of Genetic Analysis Buffer and 7.80g of water to a 20m1 glass vial. The mixture was stirred for one hour then passed through a 0.2 micron filter. Intercalating dye (Syto 61, Molecular Probes, Eugene OR) was added to the solution at a 1:2500 ratio.
Sample buffer was prepared by adding 2 ml of Gene Scan buffer to 8 nil of deionized water and 4 l of Syto 61. Separation of the 100bp ladder was again carried out under conditions and using systems identical to that described above. A representative separation is shown in Figure 4.
As can be seen from Figure 4, all 11 fragments of the ladder were separated in less than 90 seconds.
D. Sioagion of a IOObp Ladder with PolM #4 A 1.8% solution of Polymer 4 was prepared by adding 0.18g of Polymer 4, 2.Og of Genetic Analysis Buffer and 7.82g of water to a 20m1 glass vial. The mixture was stirred for one hour then passed through a 0.2 micron filter. Intercalating dye (Syto 61, Molecular Probes, Eugene OR) was added to the solution at a 1:2500 ratio.
Sample buffer was prepared by adding 2m1 of Gene Scan buffer to 8 ml of deionized water and 4 l of Syto 61. Again, a representative separation using Polymer #4 is shown in Figure 5, wherein all 11 fragments are again clearly resolved in less than 90 seconds.
As can be seen from the foregoing examples, the negatively charged polymer solutions (Polymers #2, #3, and #4) provide very high-resolution separation of the nucleic acid fragments in repeated runs. These separations were on par with, and in some cases, better than the separations obtained using neutral polymer solutions (Polymer #1).

E. Co 'g?arison of Electroosmotic Flow in Nwtral and Ne 'velv ChffLed Polvmer Solutions The level of electroosmotic flow was also determined for the neutral and charged polymer solutions (Polymers #1 and #2, respectively). The same protocol described above was used for this measurement, with the exception that Rhodamine B, a neutral fluorescent indicator of electroosmotic flow, was added to the sample buffer at 1 M
concentration, in place of the 100 bp ladder. A field of 350 mV/cm was applied to the sample well containing the Rhodamine B and its progression was visually monitored on a fluorescent detection microscope. An electroosmotic flow of 8.0 X 10-6cm2V"'s"' was measurai for the neutral, uncharged polymer solution, while an electroosmotic flow rate of 4.2 X 10-6cm2V"'s"' was measured for the negatively charged polymer solution.
Thus, in addition to providing a more than adequate reduction in electroosmotic flow, the negatively charged polymer surprisingly caused a greater reduction in that flow over neutrally charged polymer in the experiment performed. In any event, both values represent an approximate 20-fold reduction in electroosmotic flow over uncoated silica capillaries.
F. Macromolecujar SeRarations in Planar Poly,Mgnc Substrates 1. Fabrication of a Plastic Planar Ca Rillarv Structure The layout of the planar capillary structure is found in Figure 6. The, first fabrication step was to laser ablate the capillaries into a 0.2 x 3.7 x 2.2 cm piece of cast polymethylmethacrylate with an eximer laser to create a channel plate. The laser-ablated channels were measumd at 12.5 microns deep and 85 microns wide. A top plate with the same exterior dimensions as the channel plate but having 0.25-cm holes drilled through it that aligned with where the channels terminate on the channel plate was also fabricated.
The two plates were sandwiched together and then bonded by applying 10 kilograms of weight and then heating the assembly to 92C for two hours. The weight was then removed and the part was cooled to ambient temperature.
2. SUMssion of Electroosmotic Flow by Polymer #3 in a plastic Planar Canillarv The electroosmotic flow of the structure was measured first with buffer as a control. Genetic Analysis Buffer (Perkin-Elmer) was used for the measurement.
It was prepared by mixing 1 ml of the lOX buffer concentrate with 9 nil of deionized water. A
neutral dye, Bodipy-Fluorescein (Molecular Probes) was added to an aliquot of the buffer to serve as an electroosmotic flow marker. The plastic capillary structure was first filled with the buffer solution. One of the wells was then filled with the buffer containing the neutral dye. The structure was run on the microscope system described in Example E and the migration of the dye was followed visually. The electroosmotic flow was measured at 2.26 x 10-4 cm2/sec-V.

The electroosmotic flow was then measured with a 2.0% solution of Polymer #3 in Genetic Analysis Buffer. This solution was prepared as described above with the exception that Syto 61 was not added. The polymeric substrate previously described was filled with the polymer solution. One well was filled with the Bodipy Fluorescein buffer solution. The field was applied and the rate of migration of the dye was measured visually.
The field was then reversed and the migration of the dye was measured visually. With the field reversed the dye migrated at the same rate in the opposite direction.
The electroosmotic flow was calculated to be 2.54 x 10r6 cm2/sec-V, a factor of 20 lower then in the buffer control.
3. DNA SeparA ions in a Polvme_rc Device Using Poly= as a Scpara;ion Matrix All buffers and DNA samples were prepartd as described above. The microscale channel device used was described in Fl, above. The device was run on a fluorescent detection microscope system equipped with a 3mW red solid state laser (Coherent). The field applied was 210v/cm with a separation length of 1.8 cm from injection to detection point. The separation is illustrated in Figure 7 from which it can be seen that the 11 fragments in the ladder are distinguishable.

III. Mixed Poly= &parations Two different solutions of polydimethylacrylamide%oacrylic acid polymer were obtained from Polysciences, Inc. Each polymer solution had the same prepared level of negative charge (0.196). The first polymer solution was a low molecular weight polymer having an average molecular weight of approximately 100 kD. The second polymer solution was a higher molecular weight polymer solution having an average molecular weight of approximately 500 kD.
A first separation was performed on a DNA standard ladder of fragments between 50 and 100 base pairs, using only the high molecular weight polymer.
The conditions of separation were substantially as described previously. Figure 8A
shows a chromatogram of the separation of the various fragments using only this high molecular weight polymer solution at 0.8% total polymer. As can be seen, even though the peaks are well resolved, the smallest fragments (far left) are grouped closely together, i.e. less resolved. These results were also seen at 1% and 1.2% total polymer.
In a second experiment, the same DNA standard ladder was separated using a mixture of low and high molecular weight polymer. Specifically, a mixture of high molecular weight polymer (0.8%) and low molecular weight polymer (0.5%) was prepared (1.3% total polymer concentration). The chromatogram for the separation of the DNA
ladder is shown in Figure 8B. As can be seen from the chromatogram, the mixed polymer solution provides better resolution of the lower molecular.weight species, e.g., the 50 and 100 base pair fragments. While a minor loss in resolution was seen in the higher molecular weight fragments, this loss was minimal.
While the foregoing invention has been described in some detail for purposes of clarity and understanding, it will be clear to one skilled in the art from a reading of this disclosure that various changes in form and detail can be made without departing from the true scope of the invention.

Claims (155)

What is claimed is:
1. A method of separating macromolecules by capillary electrophoresis, comprising:
providing a substrate comprising at least a first capillary channel disposed therein, a surface of the channel having a first surface charge associated therewith;
filling said capillary channel with a water soluble hydrophilic polymer solution having a percent charge of from about 0.01% to about 2%, as calculated by molar percent of charged monomer subunits to total monomer utilized in producing the polymer, the charged monomer subunits consist of monomer subunits having a charge that is the same as the first surface charge;
introducing a sample containing the macromolecules into one end of the capillary channel and;
applying a voltage gradient across the length of the capillary channel, whereby the macromolecules in the sample are separated in the capillary channel.
2. The method of claim 1, wherein the substrate provided in the providing step comprises the first surface charge that is negative, and the charged monomer subunits in the filling step consist of negatively charged monomer subunits.
3. The method of claim 2, wherein the negatively charged monomer units are selected from acrylic acid, bisacrylamidoacetic acid, 4,4-Bis(4-hydroxyphenyl)pentanoic acid, 3-butene-1,2,3-tricarboxylic acid, 2-carboxyethylacrylate, itaconic acid, methacrylic acid, 4-vinylbenzoic acid, sulfonic acid, 2-acrylamido-2-methyl-1-propanesulfonic acid, 2-methyl-2-propene-1-sulfonic acid, 2-propene-1-sulfonic acid, 4-styrenesulfonic acid, 2-sulfoethyl methacrylate, 3-sulfopropyldimethyl-3-methacrylamidopropylammonium inner salt, 3-sulfopropyl methacrylate, vinylsulfonic acid, Bis(2-methacryloxyethyl)phosphate, and monoacryloxyethyl phosphate.
4. The method of claim 1, 2, or 3, wherein the substrate provided in the providing step is a silica-based substrate.
5. The method of claim 1, 2, or 3, wherein the substrate provided in the providing step comprises a solid polymeric substrate.
6. The method of claim 5, wherein the solid polymeric substrate is selected from the group of polydimethylsiloxanes (PDMS), polymethylmethacrylate (PMMA), polyurethane, polyvinylchloride (PVC), polystyrene, polysulfone, polycarbonate, polytetrafluoroethylene.
7. The method of claim 4, wherein the substrate provided in the providing step comprises a silica substrate, and the polymer in the filling step comprises polydimethylacrylamide-co-acrylic acid.
8. The method of any one of claims 1 to 7, wherein the sample contains a plurality of different nucleic acid sequences.
9. The method of claim 8, wherein the different nucleic acids comprise a plurality of different fragments of a target nucleic acid sequence.
10. The method of claim 9, wherein the different nucleic acids comprise a nested set of fragments of the target nucleic acid sequence.
11. The method of claim 10, wherein the each fragment in the nested set of fragments differs from at least one other fragment in the nested set by the addition or omission of a single nucleotide at a terminus of the fragment.
12. The method of any one of claims 1 to 11, wherein the capillary channel provided in the providing step intersects and is fluidly connected with at least a second capillary channel disposed in the substrate.
13. The method of any one of claims 1 to 11, wherein the capillary channel provided in the providing step intersects and is fluidly connected with at least second and third capillary channels disposed in the substrate.
14. The method of claim 13, wherein the applying step comprises simultaneously applying the voltage gradient across each of the first and second capillary channels, to transport the sample from the second channel into the first channel and to separate macromolecules in the sample in the first channel.
15. The method of claim 13, wherein the applying step comprises simultaneously applying the voltage gradient across each of the first, second and third capillary channels.
16. The method of any one of claims 1 to 15, wherein the polymer in the polymer solution has a net charge of between about 0.01 % and 1 %.
17. The method of any one of claims 1 to 15, wherein the polymer in the polymer solution has a net charge of between about 0.01 % and 0.5%.
18. The method of any one of claims 1 to 15, wherein the polymer in the polymer solution has a net charge of between about 0.05 % and 0.2 %.
19. The method of any one of claims 1 to 18, wherein the polymer solution comprises a polymer concentration of between about 0.01 % and about 20% (w/v).
20. The method of any one of claims 1 to 18, wherein the polymer solution comprises a polymer concentration of between about 0.1 % and about 10% (w/v).
21. The method of any one of claims 1 to 20, wherein the polymer solution has a viscosity of between about 2 centipoise and about 1000 centipoise.
22. The method of any one of claims 1 to 20, wherein the polymer solution has a viscosity in a range of from about 5 centipoise to about 200 centipoise.
23. The method of any one of claims 1 to 20, wherein the polymer solution comprises a viscosity in a range of from about 10 centipoise to about centipoise.
24. The method of any one of claims 1 to 23, wherein the polymer comprises a molecular weight from about 1 kD, to about 5,000 kD.
25. The method of any one of claims 1 to 24, wherein the polymer is a polydimethylacrylamide polymer and the charged monomer is acrylic acid.
26. The method of any one of claims 1 to 25, wherein the polymer comprises a mixture of low molecular weight polymer and high molecular weight polymer, the low molecular weight polymer having a molecular weight between about 50 kD
and about 300 kD and the high molecular weight polymer having an average molecular weight of between about 300 kD and about 1,000 kD.
27. The method of any one of claims 1 to 25, wherein the polymer comprises a mixture of low molecular weight polymer and high molecular weight polymer, the low molecular weight polymer having a molecular weight between about 50 kD
and about 200 kD and the high molecular weight polymer having an average molecular weight of between about 400 kD and about 600 kD.
28. The method of claim 26 or 27, wherein the mixture of low molecular weight polymer and high molecular weight polymer comprises a ratio of low molecular weight polymer to high molecular weight polymer of between about 1:10 and 10:1.
29. The method of claim 26 or 27, wherein the mixture of low molecular weight polymer and high molecular weight polymer comprises a ratio of low molecular weight polymer to high molecular weight polymer of between about 1:5 and 5:1.
30. The method of claim 26 or 27, wherein the mixture of low molecular weight polymer and high molecular weight polymer comprises a ratio of low molecular weight polymer to high molecular weight polymer of between about 1:2 and 2:1.
31. A method of separating macromolecules by capillary electrophoresis, comprising:
providing a silica substrate having a capillary channel disposed therein, a surface of the channel having a negative surface charge associated therewith;
filling said capillary channel with a water soluble hydrophilic polymer solution having a net charge of from about 0.1% to about 2%, the charge being the same as the surface charge;

introducing a sample containing the macromolecules into one end of the capillary channel; and applying a voltage gradient across the length of the capillary channel, whereby the macromolecules in the sample are separated in the capillary channel.
32. A method of preparing a walled capillary channel for use in separating macromolecules, comprising:
filling the walled capillary channel with a silica adsorbing polymer solution, wherein the polymer has a net charge that is the same as a net charge associated with interior surfaces of the walled capillary channel.
33. A system for separating macromolecules by capillary electrophoresis, comprising:
a substrate having at least a first walled capillary channel disposed therein, the channel having a net surface charge associated with interior surfaces of the channel;
a solution of silica adsorbing polymer disposed in the capillary channel, the solution of polymer comprising:
a molecular weight between about 1 kD and 5,000 kD;
a net charge of between about 0.01 and 2%, the net charge being the same as the net surface charge; and a power source electrically coupled to the first capillary channel for applying a voltage gradient across the capillary channel.
34. The system of claim 33, wherein the net surface charge associated with the interior surfaces of the capillary channel is negative.
35. The system of claim 34, wherein the substrate is a silica substrate.
36. The system of claim 35, wherein the substrate is selected from a silica capillary tube and an etched planar silica substrate.
37. The system of claim 33, wherein the substrate comprises a solid polymeric substrate.
38. The system of claim 37, wherein the solid polymeric substrate is selected from the group of polydimethylsiloxanes (PDMS), polymethylmethacrylate (PMMA), polyurethane, polyvinylchloride (PVC), polystyrene, polysulfone, polycarbonate, polytetrafluoroethylene.
39. The system of any one of claims 33 to 38, wherein the substrate further comprises at least a second walled capillary channel disposed in the substrate, the second walled capillary channel intersecting and in fluid communication with the first walled capillary channel.
40. The system of claim 39, wherein the power source is electrically coupled to each of the first and second capillary channels, the power source simultaneously applying the voltage gradient across a length of each of the first and second capillary channels.
41. The system of any one of claims 33 to 40, wherein the polymer has a net charge between about 0.01 % and about 1 %.
42. The system of any one of claims 33 to 40, wherein the polymer has a net charge between about 0.01 % and 0.5 %.
43. The system of any one of claims 33 to 40, wherein the polymer has a net charge between about 0.05 % and 0.2 %.
44. The system of any one of claims 33 to 43, wherein the polymer solution comprises a polymer concentration in a range of from about 0.01 % to about 20 %
(w/v).
45. The system of any one of claims 33 to 43, wherein the polymer solution comprises a polymer concentration in a range of from about 0.1 % to about 10 %
(w/v).
46. The system of any one of claims 33 to 45, wherein the polymer solution comprises a viscosity of between about 2 centipoise and about 1000 centipoise.
47. The system of any one of claims 33 to 45, wherein the polymer solution comprises a viscosity in a range of from about 5 centipoise to about 200 centipoise.
48. The system of any one of claims 33 to 45, wherein the polymer solution comprises a viscosity in a range of from about 10 centipoise to about centipoise.
49. The system of any one of claims 33 to 48, wherein the polymer is an acrylic polymer and the charged monomer subunits are selected from acrylic acid, bisacrylamidoacetic acid, 4,4-Bis(4-hydroxyphenyl)pentanoic acid, 3-butene-1,2,3-tricarboxylic acid, 2-carboxyethylacrylate, itaconic acid, methacrylic acid, 4-vinylbenzoic acid, sulfonic acid, 2-acrylamido-2-methyl-1-propanesulfonic acid, 2-methyl-2-propene-1-sulfonic acid, 2-propene-1-sulfonic acid, 4-styrenesulfonic acid, 2-sulfoethyl methacrylate, 3-sulfopropyldimethyl-3-methacrylamidopropylammonium inner salt, 3-sulfopropyl methacrylate, vinylsulfonic acid, Bis(2-methacryloxyethyl)phosphate, and monoacryloxyethyl phosphate.
50. The system of any one of claims 33 to 48, wherein the polymer comprises polydimethylacrylamide-co-acrylic acid.
51. The system of any one of claims 33 to 48, wherein the polymer has a net negative charge.
52. The system of any one of claims 33 to 51, wherein the polymer is made by the process of polymerizing dimethylacrylamide monomers in the presence acrylic acid, the acrylic acid being present at a concentration of between about 0.01 and 2 % of a total monomer concentration.
53. The system of any one of claims 33 to 52, wherein the first net surface charge is capable of supporting an electroosmotic mobility of a buffer comprising from about 1 mM to about 10 mM sodium borate buffer, at a pH of from about 7 to about 9, disposed in the walled capillary channel, the electroosmotic mobility being at least about 1 X 10 -5cm2V-1s-1.
54. A system for separating nucleic acids by molecular weight, comprising:
a silica substrate having a walled capillary channel disposed therein, the channel having a negative charge associated with interior surfaces of the channel;
a solution of silica adsorbing polymer disposed in the capillary channel, the solution of polymer comprising:
a molecular weight between about 1 kD and 5,000 kD;
a net negative charge of between about 0.01 and 2%; and a power source for applying a voltage gradient across the capillary channel.
55. A walled capillary for separating macromolecules by capillary electrophoresis, comprising:
a capillary channel disposed in a solid substrate, interior surfaces of the capillary channel having a first net surface charge associated therewith; and a solution of silica adsorbing polymer disposed in the capillary channel, the polymer comprising:
a molecular weight between about 1 kD and about 5,000 kD;
a net charge of between about 0.01 and 2%, the net charge being the same as the first net surface charge.
56. A method of manufacturing a microfabricated channel system, the method comprising:
providing a device comprising at least one microchannel; and, disposing a polymer in the at least one microchannel, the polymer comprising a net charge of between about 0.01% and 2%, the net charge being of the same charge as at least one surface of the microchannel.
57. The method of claim 56, wherein the polymer has a net charge of between about 0.01% and about 1%.
58. The method of claim 56, wherein the polymer has a net charge of between about 0.01% and about 0.5%.
59. The method of claim 56, wherein the polymer has a net charge of between about 0.05% and 0.5%.
60. The method of claim 56, wherein the polymer has a net charge of between about 0.05% and 0.2%.
61. The method of any one of claims 56 to 60, wherein the polymer has a molecular weight between about 1 Kd and about 5,000 Kd.
62. The method of any one of claims 56 to 60, wherein the polymer has a molecular weight between about 1 Kd and about 6,000 Kd.
63. The method of any one of claims 56 to 60, wherein the polymer has a molecular weight between about 1 Kd about 1000 Kd.
64. The method of any one of claims 56 to 63, wherein the polymer is a water soluble polymer.
65. The method of any one of claims 56 to 63, wherein the polymer is a water soluble surface adsorbing polymer.
66. The method of any one of claims 56 to 63, wherein the polymer is a water soluble silica adsorbing polymer.
67. The method of any one of claims 56 to 63, wherein the polymer is a water soluble surface adsorbing acrylic polymer.
68. The method of any one of claims 56 to 63, wherein the polymer is a water soluble surface adsorbing polydimethylacrylamide polymer-co-acrylic acid polymer.
69. The method of any one of claims 56 to 63, wherein the polymer is a water soluble surface adsorbing dimethylacrylamide polymer incorporating a selected percentage of charged acrylic acid monomers.
70. The method of any one of claims 56 to 63, wherein the polymer is a water soluble surface adsorbing polymer which polymer is an acrylic polymer selected from a polyacrylamide, a polymethylacrylamide, and a polydimethylacrylamide.
71. The method of any one of claims 56 to 63, wherein the polymer is a water soluble surface adsorbing polymer formed from a carboxylic acid monomer.
72. The method of any one of claims 56 to 63, wherein the polymer is a water soluble surface adsorbing polymer formed from a monomer selected from:
acrylic acid, bisacrylamidoacetic acid, 4,4-Bis(4-hydroxyphenyl)pentanoic acid, 3-butene-1,2,3-tricarboxylic acid, 2-carboxyethylacrylate, itaconic acid, methacrylic acid, and 4-vinylbenzoic acid.
73. The method of any one of claims 56 to 63, wherein the polymer is a water soluble surface adsorbing polymer formed from a sulfonic acid or phosphoric acid monomer.
74. The method of any one of claims 56 to 63, wherein the polymer is a water soluble surface adsorbing polymer formed from a sulfonic acid or phosphoric acid monomer selected from 2-acrylamido-2-methyl-1-propanesulfonic acid, 2-methyl-2-propene-1-sulfonic acid, 2-propene-1-sulfonic acid, 4-styrenesulfonic acid, 2-sulfoethyl methacrylate, 3-sulfopropyldimethyl-3-methacrylamidopropylammonium inner salt, sulfopropyl methacrylate, vinylsulfonic acid, Bis(2-methacryloxyethyl)phosphate, and monoacryloxyethyl phosphate.
75. The method of any one of claims 56 to 74, wherein the surface of the microchannel comprises a first surface which is negatively charged and wherein monomer subunits of the polymer comprise negatively charged monomer subunits.
76. The method of claim 75, wherein the negatively charged monomer units are selected from: acrylic acid, bisacrylamidoacetic acid, 4,4-Bis(4-hydroxyphenyl)pentanoic acid, 3-butene-1,2,3-tricarboxylic acid, 2-carboxyethylacrylate, itaconic acid, methacrylic acid, 4-vinylbenzoic acid, sulfonic acid, 2-acrylamido-2-methyl-1-propanesulfonic acid, 2-methyl-2-propene1l-sulfonic acid, 2-propene-1-sulfonic acid, 4-styrenesulfonic acid, 2-sulfoethyl methacrylate, 3-sulfopropyldimethyl-3-methacrylamidopropylammonium inner salt, 3-sulfopropyl methacrylate, vinylsulfonic acid, Bis(2-methacryloxyethyl)phosphate, and monoacryloxyethyl phosphate.
77. The method of any one of claims 56 to 74, wherein the microchannel comprises a region of positive charge and the polymer is formed from positively charged monomer units.
78. The method of any one of claims 56 to 74, wherein the microchannel comprises a region of positive charge and the polymer is formed from positively charged monomer units, which monomer units are quaternary amine monomers.
79. The method of any one of claims 56 to 74, wherein the microchannel comprises a region of positive charge and the polymer is formed from positively charged monomer units, which monomer units are quaternary amine monomers selected from acryloxyethyltrimethylammonium chloride, diallyldimethylammonium chloride, 2-methacryloxyethyltrimethylammonium chloride, and 3-methacryloxy-2-hydroxypropyltrimethylammonium chloride.
80. The method of any one of claims 56 to 79, wherein the polymer is provided as a water soluble hydrophilic polymer solution and the method further comprising filling the microchannel with the water soluble hydrophilic polymer solution.
81. The method of any one of claims 56 to 79, wherein the polymer is provided in an aqueous solution at a concentration between about 0.01 % and 30% (w/v).
82. The method of any one of claims 56 to 79, wherein the polymer is provided in an aqueous solution at a concentration between about 0.01 % and about 20%
(w/v).
83. The method of any one of claims 56 to 79, wherein the polymer is provided in an aqueous solution at a concentration between about 0.1 % and about 10%
(w/v).
84. The method of any one of claims 56 to 79, wherein the polymer is in a solution which has a solution viscosity, as used within the microchannel, of from about 2 to about 1000 centipoise.
85. The method of any one of claims 56 to 79, wherein the polymer is in a solution which has a solution viscosity, as used within the microchannel, of from about 5 to about 200 centipoise.
86. The method of any one of claims 56 to 79, wherein the polymer is in a solution which has a solution viscosity, as used within the microchannel, of from about 10 to about 100 centipoise.
87. The method of any one of claims 56 to 79, wherein the polymer is in a solution which has a solution viscosity as measured at 20° C using an Ubberholde viscometer of about 5.7, 7.4, 34.2, or 60.1 centipoise.
88. The method of any one of claims 56 to 79, wherein the polymer is present in a solution which further comprises a buffer for controlling pH or conductivity, or wherein the solution further comprises an additional polymer.
89. The method of any one of claims 56 to 88, wherein the microchannel further comprises a neutral polymer.
90. The method of any one of claims 56 to 88, wherein the microchannel further comprises a neutral polymer, which neutral polymer enhances sieving of an analyte in the microchannel.
91. The method of any one of claims 56 to 63, wherein the polymer is formed by 2-Methyl-1-propanol polymerization of polydimethylacrylamide.
92. The method of any one of claims 56 to 63, wherein the polymer is formed by 2-Methyl-1-propanol polymerization of polydimethylacrylamide/acrylic acid.
93. The method of any one of claims 56 to 63, wherein the polymer is formed by aqueous polymerization of medium molecular weight polydimethylacryl-amide/acrylic acid.
94. The method of any one of claims 56 to 63, wherein the polymer is formed by aqueous polymerization of high molecular weight polydimethylacryl-amide/acrylic acid.
95. The method of any one of claims 56 to 79 and 91 to 94, wherein the polymer is disposed in the microchannel by placing one end of the channel into contact with a solution comprising the polymer and allowing the polymer to wick into the channel.
96. The method of any one of claims 56 to 94, wherein the polymer is disposed in the microchannel by driving the polymer into the channel using vacuum or pressure.
97. The method of any one of claims 56 to 79 and 91 to 94, wherein the polymer is disposed in the microchannel by placing the microchannel in contact with a polymer solution comprising the polymer at a reservoir comprising the polymer solution disposed at the end of the channel, and applying a slight pressure to force the polymer into the microchannel.
98. The method of any one of claims 56 to 97, further comprising adding DNA to the microchannel.
99. The method of any one of claims 56 to 98, wherein the device is a planar microfabricated structure comprising a plurality of microscale capillary channels fabricated therein.
100. The method of claim 99, wherein the planar microfabricated structure is provided by fabricating a channel network into the surface of a planar substrate and overlaying a second substrate on the surface of the first substrate to cover and seal the channels, thereby providing the microscale capillary channels.
101. The method of any one of claims 56 to 100, wherein the microchannel has a charged surface capable of supporting an electroosmotic mobility (µEO) of at least about 1 × 10-5 cm2 V-1 s-1, for a buffer when said buffer is in contact with said microchannel, which buffer comprises from about 1 mM to about 100 mM sodium borate at a pH of from about 6 to about 9.
102. The method of any one of claims 56 to 101, wherein the microchannel has an internal cross-sectional dimension of between about 1 µm and about 500 µm.
103. The method of any one of claims 56 to 74, wherein the at least one microchannel comprises a material selected from: silica, polydimethylsiloxanes (PDMS), polymethylmethacrylate (PMMA), polyurethane, polyvinylchloride (PVC), polystyrene, polysulfone, polycarbonate, polytetrafluoroethylene.
104. A microfabricated channel system made by the method of any one of claims 56 to 103.
105. A device, comprising: at least one microchannel, the microchannel comprising a polymer disposed therein, the polymer comprising a net charge of between about 0.01 and 2%, the net charge being of the same charge as at least one surface of the microchannel.
106. The device of claim 105, wherein the polymer has a net charge of between about 0.01 % and about 1%.
107. The device of claim 105, wherein the polymer has a net charge of between about 0.01 % and about 0.5%.
108. The device of claim 105, wherein the polymer has a net charge of between about 0.05% and 0.5%.
109. The device of claim 105, wherein the polymer has a net charge of between about 0.05% and 0.2%.
110. The device of any one of claims 105 to 109, wherein the polymer has a molecular weight between about 1 Kd and about 5,000 Kd.
111. The device of any one of claims 105 to 109, wherein the polymer has a molecular weight between about 1 Kd and about 6,000 Kd.
112. The device of any one of claims 105 to 109, wherein the polymer has a molecular weight between about 1 Kd about 1000 Kd.
113. The device of any one of claims 105 to 112, wherein the polymer is a water soluble polymer.
114. The device of any one of claims 105 to 112, wherein the polymer is a water soluble surface adsorbing polymer.
115. The device of any one of claims 105 to 112, wherein the polymer is a water soluble silica adsorbing polymer.
116. The device of any one of claims 105 to 112, wherein the polymer is a water soluble surface adsorbing acrylic polymer.
117. The device of any one of claims 104 to 112, wherein the polymer is a water soluble surface adsorbing polydimethylacrylamide polymer-co-acrylic acid polymer.
118. The device of any one of claims 104 to 112, wherein the polymer is a water soluble surface adsorbing dimethylacrylamide polymer incorporating a selected percentage of charged acrylic acid monomers.
119. The device of any one of claims 104 to 112, wherein the polymer is a water soluble surface adsorbing acrylic polymer selected from a polyacrylamide, a polymethylacrylamide, and a polydimethylacrylamide.
120. The device of any one of claims 104 to 112, wherein the polymer is a water soluble surface adsorbing polymer formed from a carboxylic acid monomer.
121. The device of any one of claims 104 to 112, wherein the polymer is a water soluble surface adsorbing polymer is formed from a monomer selected from: acrylic acid, bisacrylamidoacetic acid, 4,4-Bis(4-hydroxyphenyl)pentanoic acid, 3-butene-1,2,3-tricarboxylic acid, 2-carboxyethylacrylate, itaconic acid, methacrylic acid, and 4-vinylbenzoic acid.
122. The device of any one of claims 104 to 112, wherein the polymer is a water soluble surface adsorbing polymer formed from a sulfonic acid or phosphoric acid monomer.
123. The device of any one of claims 104 to 112, wherein the polymer is a water soluble surface adsorbing polymer formed from a sulfonic acid or phosphoric acid monomer selected from 2-acrylamido-2-methyl-1-propanesulfonic acid, 2-methyl-2-propene-1-sulfonic acid, 2-propene-1-sulfonic acid, 4-styrenesulfonic acid, 2-sulfoethyl methacrylate, 3-sulfopropyldimethyl-3-methacrylamidopropylammonium inner salt, sulfopropyl methacrylate, vinylsulfonic acid, Bis(2-methacryloxyethyl)phosphate, and monoacryloxyethyl phosphate.
124. The device of any one of claims 104 to 123, wherein the surface of the microchannel comprises a first surface which is negatively charged and monomer subunits of the polymer comprise negatively charged monomer subunits.
125. The device of claim 124, wherein the negatively charged monomer units are selected from: acrylic acid, bisacrylamidoacetic acid, 4,4-Bis(4-hydroxyphenyl) pentanoic acid, 3-butene-1,2,3-tricarboxylic acid, 2-carboxyethylacrylate, itaconic acid, methacrylic acid, 4-vinylbenzoic acid, sulfonic acid, 2-acrylamido-2-methyl-1-propanesulfonic acid, 2-methyl-2-propene-1-sulfonic acid, 2-propene-1-sulfonic acid, 4-styrenesulfonic acid, 2-sulfoethyl methacrylate, 3-sulfopropyldimethyl-3-methacrylamidopropylammonium inner salt, 3-sulfopropyl methacrylate, vinylsulfonic acid, Bis(2-methacryloxyethyl)phosphate, and monoacryloxyethyl phosphate.
126. The device of any one of claims 104 to 123, wherein the microchannel comprises a region of positive charge and the polymer is formed from positively charged monomer units.
127. The device of any one of claims 104 to 123, wherein the microchannel comprises a region of positive charge and the polymer is formed from positively charged monomer units, which monomer units are quaternary amine monomers.
128. The device of any one of claims 104 to 123, wherein the microchannel comprises a region of positive charge and the polymer is formed from positively charged monomer units, which monomer units are quatemary amine monomers selected from 2-acryloxyethyltrimethylammonium chloride, diallyldimethylammonium chloride, 2-methacryloxyethyltrimethylammonium chloride, and 3-methacryloxy-2-hydroxypropyltrimethylammonium chloride.
129. The device of any one of claims 104 to 128, wherein the microchannel is filled with a water soluble hydrophilic polymer solution.
130. The device of any one of claims 104 to 128, wherein the polymer is in an aqueous solution at a concentration between about 0.01 % and 30% (w/v).
131. The device of any one of claims 104 to 128, wherein the polymer is in an aqueous solution at a concentration between about 0.01 % and about 20%
(w/v).
132. The device of any one of claims 104 to 128, wherein the polymer is in an aqueous solution at a concentration between about 0.1 % and about 10%
(w/v).
133. The device of any one of claims 104 to 128, wherein the polymer is in a solution which has a solution viscosity, within the microchannel, of from about 2 to about 1000 centipoise.
134. The device of any one of claims 104 to 128, wherein the polymer is in a solution which has a solution viscosity, within the microchannel, of from about 5 to about 200 centipoise.
135. The device of any one of claims 104 to 128, wherein the polymer is in a solution which has a solution viscosity, within the microchannel, of from about 10 to about 100 centipoise.
136. The device of any one of claims 104 to 128, wherein the polymer is in a solution which has a solution viscosity, as measured at 20° C
using an Ubberholde viscometer, of about 5.7, 7.4, 34.2, or 60.1 centipoise.
137. The device of any one of claims 104 to 128, wherein the polymer is in a solution which further comprises a buffer for controlling pH or conductivity, or wherein the solution comprises one or more additional polymers.
138. The device of any one of claims 104 to 137, wherein the microchannel further comprises a neutral polymer.
139. The device of any one of claims 104 to 137, wherein the microchannel further comprises a neutral polymer, which neutral polymer enhances sieving of an analyte in the microchannel.
140. The device of any one of claims 104 to 112, wherein the polymer is formed by 2-Methyl-1-propanol polymerization of polydimethylacrylamide.
141. The device of any one of claims 104 to 112, wherein the polymer is formed by 2-Methyl-1-propanol polymerization of polydimethylacrylamide/acrylic acid.
142. The device of any one of claims 104 to 112, wherein the polymer is formed by aqueous polymerization of medium molecular weight polydimethylacrylamide/acrylic acid.
143. The device of any one of claims 104 to 112, wherein the polymer is formed by aqueous polymerization of high molecular weight polydimethylacrylamide/acrylic acid.
144. The device of any one of claims 104 to 128 and 140 to 143, wherein the polymer is disposed in the microchannel by placing one end of the channel into contact with a solution comprising the polymer and allowing the polymer to wick into the channel.
145. The device of any one of claims 104 to 143, wherein the polymer is disposed in the microchannel by driving the polymer into the channel using vacuum or pressure.
146. The device of any one of claims 104 to 128 and 140 to 143, wherein the polymer is disposed in the microchannel by placing the microchannel in contact with a polymer solution comprising the polymer at a reservoir comprising the polymer solution disposed at the end of the channel, and applying a slight pressure to force the polymer into the microchannel.
147. The device of any one of claims 104 to 146, further comprising DNA
in the microchannel.
148. The device of any one of claims 104 to 147, wherein the device comprises a planar microfabricated structure comprising a plurality of microscale capillary channels fabricated therein, wherein the at least one microchannel is one of the plurality of microscale capillary channels.
149. The device of claim 148, wherein the planar microfabricated structure is provided by fabricating a channel network into the surface of a planar substrate and overlaying a second substrate on the surface of the first substrate to cover and seal the channels, thereby providing the microscale capillary channels.
150. The device of any one of claims 104 to 149, wherein the microchannel has a charged surface capable of supporting an electroosmotic mobility (µEO) of at least about 1 × 10-5 cm2 V-1 s-1, for a buffer when that buffer is in contact with those walls, which buffer comprises from about 1 mM to about 100 mM sodium borate at a pH of from about 6 to about 9.
151. The device of any one of claims 104 to 150, wherein the microchannel has an internal cross-sectional dimension of between about 1 µm and about 500 µm.
152. The device of any one of claims 104 to 123, wherein the at least one microchannel comprises a material selected from: silica, polydimethylsiloxanes (PDMS), polymethylmethacrylate (PMMA), polyurethane, polyvinylchloride (PVC), polystyrene, polysulfone, polycarbonate, polytetrafluoroethylene.
153. The device of any one of claims 104 to 152, further comprising an electrode in fluidic contact with the polymer in the microscale channel.
154. The device of any one of claims 104 to 152, further comprising a plurality of electrodes in fluidic contact with the polymer in the microscale channel, which electrodes during operation of the device, apply an electric field along the microscale channel, thereby providing for electrophoresis of analytes through the microscale channel.
155. The device of any one of claims 104 to 154, further comprising at least one well or reservoir in fluidic contact with the microscale channel, which well or reservoir is coupled to a pressure source, which pressure source directs flow of the polymer into the channel.
CA002309831A 1997-12-17 1998-12-02 Improved methods and systems for performing molecular separations Expired - Fee Related CA2309831C (en)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
US08/992,239 1997-12-17
US08/992,239 US5948227A (en) 1997-12-17 1997-12-17 Methods and systems for performing electrophoretic molecular separations
PCT/US1998/025562 WO1999031495A1 (en) 1997-12-17 1998-12-02 Improved methods and systems for performing molecular separations

Publications (2)

Publication Number Publication Date
CA2309831A1 CA2309831A1 (en) 1999-06-24
CA2309831C true CA2309831C (en) 2007-07-17

Family

ID=25538087

Family Applications (1)

Application Number Title Priority Date Filing Date
CA002309831A Expired - Fee Related CA2309831C (en) 1997-12-17 1998-12-02 Improved methods and systems for performing molecular separations

Country Status (6)

Country Link
US (4) US5948227A (en)
EP (1) EP1040343A4 (en)
JP (1) JP4144776B2 (en)
AU (1) AU747232B2 (en)
CA (1) CA2309831C (en)
WO (1) WO1999031495A1 (en)

Families Citing this family (276)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6048734A (en) 1995-09-15 2000-04-11 The Regents Of The University Of Michigan Thermal microvalves in a fluid flow method
US5942443A (en) * 1996-06-28 1999-08-24 Caliper Technologies Corporation High throughput screening assay systems in microscale fluidic devices
US5885470A (en) 1997-04-14 1999-03-23 Caliper Technologies Corporation Controlled fluid transport in microfabricated polymeric substrates
NZ333346A (en) 1996-06-28 2000-03-27 Caliper Techn Corp High-throughput screening assay systems in microscale fluidic devices
CN1105914C (en) * 1997-04-25 2003-04-16 卡钳技术有限公司 Microfluidic devices incorporating improved channel geometries
US7033474B1 (en) 1997-04-25 2006-04-25 Caliper Life Sciences, Inc. Microfluidic devices incorporating improved channel geometries
AU730827B2 (en) * 1997-06-09 2001-03-15 Caliper Technologies Corporation Apparatus and methods for correcting for variable velocity in microfluidic systems
US6425972B1 (en) 1997-06-18 2002-07-30 Calipher Technologies Corp. Methods of manufacturing microfabricated substrates
US5989402A (en) 1997-08-29 1999-11-23 Caliper Technologies Corp. Controller/detector interfaces for microfluidic systems
US6833242B2 (en) 1997-09-23 2004-12-21 California Institute Of Technology Methods for detecting and sorting polynucleotides based on size
US5948227A (en) * 1997-12-17 1999-09-07 Caliper Technologies Corp. Methods and systems for performing electrophoretic molecular separations
US6251343B1 (en) 1998-02-24 2001-06-26 Caliper Technologies Corp. Microfluidic devices and systems incorporating cover layers
US7497994B2 (en) 1998-02-24 2009-03-03 Khushroo Gandhi Microfluidic devices and systems incorporating cover layers
US6756019B1 (en) 1998-02-24 2004-06-29 Caliper Technologies Corp. Microfluidic devices and systems incorporating cover layers
US6123798A (en) * 1998-05-06 2000-09-26 Caliper Technologies Corp. Methods of fabricating polymeric structures incorporating microscale fluidic elements
CA2332919A1 (en) * 1998-06-08 1999-12-16 Caliper Technologies Corporation Microfluidic devices, systems and methods for performing integrated reactions and separations
US6117326A (en) * 1998-06-26 2000-09-12 Rohm And Haas Company Capillary electrochromatography separation media
US6540896B1 (en) 1998-08-05 2003-04-01 Caliper Technologies Corp. Open-Field serial to parallel converter
US6149787A (en) 1998-10-14 2000-11-21 Caliper Technologies Corp. External material accession systems and methods
US7115422B1 (en) * 1998-10-23 2006-10-03 Micron Technology, Inc. Separation apparatus including porous silicon column
US6762057B1 (en) * 1998-10-23 2004-07-13 Micron Technology, Inc. Separation apparatus including porous silicon column
US20020019059A1 (en) * 1999-01-28 2002-02-14 Calvin Y.H. Chow Devices, systems and methods for time domain multiplexing of reagents
EP1159605B1 (en) * 1999-02-02 2012-07-11 Caliper Life Sciences, Inc. Methods for characterizing proteins
ATE469699T1 (en) * 1999-02-23 2010-06-15 Caliper Life Sciences Inc MANIPULATION OF MICROPARTICLES IN MICROFLUID SYSTEMS
US6749814B1 (en) * 1999-03-03 2004-06-15 Symyx Technologies, Inc. Chemical processing microsystems comprising parallel flow microreactors and methods for using same
US6558945B1 (en) * 1999-03-08 2003-05-06 Aclara Biosciences, Inc. Method and device for rapid color detection
US6500323B1 (en) 1999-03-26 2002-12-31 Caliper Technologies Corp. Methods and software for designing microfluidic devices
US6306273B1 (en) * 1999-04-13 2001-10-23 Aclara Biosciences, Inc. Methods and compositions for conducting processes in microfluidic devices
US6322683B1 (en) * 1999-04-14 2001-11-27 Caliper Technologies Corp. Alignment of multicomponent microfabricated structures
WO2000070080A1 (en) 1999-05-17 2000-11-23 Caliper Technologies Corp. Focusing of microparticles in microfluidic systems
US6592821B1 (en) 1999-05-17 2003-07-15 Caliper Technologies Corp. Focusing of microparticles in microfluidic systems
US6635163B1 (en) * 1999-06-01 2003-10-21 Cornell Research Foundation, Inc. Entropic trapping and sieving of molecules
US6649358B1 (en) 1999-06-01 2003-11-18 Caliper Technologies Corp. Microscale assays and microfluidic devices for transporter, gradient induced, and binding activities
US6375818B1 (en) * 1999-06-08 2002-04-23 Beckman Coulter, Inc. Surfaces with reduced electroosmotic flow
AU6068300A (en) 1999-07-06 2001-01-22 Caliper Technologies Corporation Microfluidic systems and methods for determining modulator kinetics
US7138254B2 (en) 1999-08-02 2006-11-21 Ge Healthcare (Sv) Corp. Methods and apparatus for performing submicroliter reactions with nucleic acids or proteins
US6495104B1 (en) 1999-08-19 2002-12-17 Caliper Technologies Corp. Indicator components for microfluidic systems
US6410668B1 (en) 1999-08-21 2002-06-25 Marcella Chiari Robust polymer coating
US6858185B1 (en) * 1999-08-25 2005-02-22 Caliper Life Sciences, Inc. Dilutions in high throughput systems with a single vacuum source
US6613581B1 (en) 1999-08-26 2003-09-02 Caliper Technologies Corp. Microfluidic analytic detection assays, devices, and integrated systems
WO2001017797A1 (en) 1999-09-10 2001-03-15 Caliper Technologies Corp. Microfabrication methods and devices
US6906797B1 (en) * 1999-09-13 2005-06-14 Aclara Biosciences, Inc. Side light activated microfluid channels
JP2003511682A (en) * 1999-10-08 2003-03-25 カリパー・テクノロジーズ・コープ. Use of Nernst voltage-sensitive dyes in transmembrane voltage measurements.
US6386014B1 (en) 1999-11-18 2002-05-14 Eagle Research Corporation Energy measurement device for flowing gas using microminiature gas chromatograph
US6468761B2 (en) 2000-01-07 2002-10-22 Caliper Technologies, Corp. Microfluidic in-line labeling method for continuous-flow protease inhibition analysis
US7037416B2 (en) * 2000-01-14 2006-05-02 Caliper Life Sciences, Inc. Method for monitoring flow rate using fluorescent markers
WO2001057509A1 (en) * 2000-02-04 2001-08-09 Caliper Technologies Corp. Methods, devices, and systems for monitoring time dependent reactions
EP1261427B1 (en) 2000-03-02 2011-03-02 Microchips, Inc. Microfabricated devices and methods for storage and selective exposure of chemicals
US7485454B1 (en) * 2000-03-10 2009-02-03 Bioprocessors Corp. Microreactor
US6749735B1 (en) 2000-03-16 2004-06-15 David Le Febre Electromobility focusing controlled channel electrophoresis system
US7141152B2 (en) * 2000-03-16 2006-11-28 Le Febre David A Analyte species separation system
US20020012971A1 (en) * 2000-03-20 2002-01-31 Mehta Tammy Burd PCR compatible nucleic acid sieving medium
US6733645B1 (en) 2000-04-18 2004-05-11 Caliper Technologies Corp. Total analyte quantitation
US6787016B2 (en) * 2000-05-01 2004-09-07 Aclara Biosciences, Inc. Dynamic coating with linear polymer mixture for electrophoresis
EP1281002A4 (en) * 2000-05-11 2006-08-09 Caliper Life Sciences Inc Microfluidic devices and methods to regulate hydrodynamic and electrical resistance utilizing bulk viscosity enhancers
AU6152301A (en) 2000-05-12 2001-11-26 Caliper Techn Corp Detection of nucleic acid hybridization by fluorescence polarization
US6730072B2 (en) 2000-05-30 2004-05-04 Massachusetts Institute Of Technology Methods and devices for sealing microchip reservoir devices
US7351376B1 (en) * 2000-06-05 2008-04-01 California Institute Of Technology Integrated active flux microfluidic devices and methods
WO2002000100A2 (en) * 2000-06-23 2002-01-03 Daniel Armstrong Method for separation, identification and evaluation of microbes and cells
US6720187B2 (en) * 2000-06-28 2004-04-13 3M Innovative Properties Company Multi-format sample processing devices
US6734401B2 (en) * 2000-06-28 2004-05-11 3M Innovative Properties Company Enhanced sample processing devices, systems and methods
US7192559B2 (en) * 2000-08-03 2007-03-20 Caliper Life Sciences, Inc. Methods and devices for high throughput fluid delivery
US20020142618A1 (en) * 2000-08-04 2002-10-03 Caliper Technologies Corp. Control of operation conditions within fluidic systems
IL155179A0 (en) * 2000-10-10 2003-11-23 Genset Sa Surface adsorbing polymers and their use in treating surfaces
EP1339312B1 (en) * 2000-10-10 2006-01-04 Microchips, Inc. Microchip reservoir devices using wireless transmission of power and data
US7106818B2 (en) * 2000-10-17 2006-09-12 Koninkijke Philips Electronics N.V. Method and apparatus for timing recovery based on dispersion characterization and components therefor
US20030057092A1 (en) * 2000-10-31 2003-03-27 Caliper Technologies Corp. Microfluidic methods, devices and systems for in situ material concentration
US20050011761A1 (en) * 2000-10-31 2005-01-20 Caliper Technologies Corp. Microfluidic methods, devices and systems for in situ material concentration
ATE432466T1 (en) 2000-10-31 2009-06-15 Caliper Life Sciences Inc MICROFLUIDIC PROCESS FOR IN-SITU MATERIAL CONCENTRATION
US20090118139A1 (en) 2000-11-07 2009-05-07 Caliper Life Sciences, Inc. Microfluidic method and system for enzyme inhibition activity screening
US6942778B1 (en) 2000-11-28 2005-09-13 Nanogen, Inc. Microstructure apparatus and method for separating differently charged molecules using an applied electric field
WO2002052045A1 (en) * 2000-12-26 2002-07-04 Aviva Biosciences Active and biocompatible platforms prepared by polymerization of surface coating films
WO2002055058A2 (en) * 2001-01-09 2002-07-18 Microchips, Inc. Flexible microchip devices for ophthalmic and other applications
WO2002060754A1 (en) 2001-01-29 2002-08-08 Caliper Technologies Corp. Non-mechanical valves for fluidic systems
US6692700B2 (en) 2001-02-14 2004-02-17 Handylab, Inc. Heat-reduction methods and systems related to microfluidic devices
AU2002327165B2 (en) * 2001-02-15 2006-08-10 Caliper Life Sciences, Inc. Microfluidic systems with enhanced detection systems
US7670559B2 (en) 2001-02-15 2010-03-02 Caliper Life Sciences, Inc. Microfluidic systems with enhanced detection systems
US6720148B1 (en) 2001-02-22 2004-04-13 Caliper Life Sciences, Inc. Methods and systems for identifying nucleotides by primer extension
US7867776B2 (en) * 2001-03-02 2011-01-11 Caliper Life Sciences, Inc. Priming module for microfluidic chips
US7150999B1 (en) 2001-03-09 2006-12-19 Califer Life Sciences, Inc. Process for filling microfluidic channels
US7316769B2 (en) * 2001-03-19 2008-01-08 Cornell Research Foundation, Inc. Length-dependent recoil separation of long molecules
US7323140B2 (en) 2001-03-28 2008-01-29 Handylab, Inc. Moving microdroplets in a microfluidic device
US6575188B2 (en) 2001-07-26 2003-06-10 Handylab, Inc. Methods and systems for fluid control in microfluidic devices
US6852287B2 (en) 2001-09-12 2005-02-08 Handylab, Inc. Microfluidic devices having a reduced number of input and output connections
US7829025B2 (en) 2001-03-28 2010-11-09 Venture Lending & Leasing Iv, Inc. Systems and methods for thermal actuation of microfluidic devices
US7010391B2 (en) 2001-03-28 2006-03-07 Handylab, Inc. Methods and systems for control of microfluidic devices
US8895311B1 (en) 2001-03-28 2014-11-25 Handylab, Inc. Methods and systems for control of general purpose microfluidic devices
EP1384022A4 (en) 2001-04-06 2004-08-04 California Inst Of Techn Nucleic acid amplification utilizing microfluidic devices
US20050032204A1 (en) * 2001-04-10 2005-02-10 Bioprocessors Corp. Microreactor architecture and methods
US20040058437A1 (en) * 2001-04-10 2004-03-25 Rodgers Seth T. Materials and reactor systems having humidity and gas control
US20030077817A1 (en) * 2001-04-10 2003-04-24 Zarur Andrey J. Microfermentor device and cell based screening method
US20040132166A1 (en) * 2001-04-10 2004-07-08 Bioprocessors Corp. Determination and/or control of reactor environmental conditions
US20040058407A1 (en) * 2001-04-10 2004-03-25 Miller Scott E. Reactor systems having a light-interacting component
CA2702143C (en) 2001-06-05 2014-02-18 Mikro Systems, Inc. Methods for manufacturing three-dimensional devices and devices created thereby
US7723123B1 (en) * 2001-06-05 2010-05-25 Caliper Life Sciences, Inc. Western blot by incorporating an affinity purification zone
US7785098B1 (en) 2001-06-05 2010-08-31 Mikro Systems, Inc. Systems for large area micro mechanical systems
US7141812B2 (en) * 2002-06-05 2006-11-28 Mikro Systems, Inc. Devices, methods, and systems involving castings
US20020187564A1 (en) * 2001-06-08 2002-12-12 Caliper Technologies Corp. Microfluidic library analysis
WO2002101375A2 (en) * 2001-06-13 2002-12-19 Symyx Technologies, Inc. Polymer additive for use in separation media
US6977163B1 (en) 2001-06-13 2005-12-20 Caliper Life Sciences, Inc. Methods and systems for performing multiple reactions by interfacial mixing
ATE285756T1 (en) * 2001-06-28 2005-01-15 Microchips Inc METHOD FOR HERMETIC SEALING MICROCHIP RESERVOIR DEVICES
ATE465811T1 (en) * 2001-07-13 2010-05-15 Caliper Life Sciences Inc METHOD FOR SEPARATING COMPONENTS OF A MIXTURE
US6825127B2 (en) 2001-07-24 2004-11-30 Zarlink Semiconductor Inc. Micro-fluidic devices
US7060171B1 (en) 2001-07-31 2006-06-13 Caliper Life Sciences, Inc. Methods and systems for reducing background signal in assays
JP4657524B2 (en) * 2001-08-29 2011-03-23 嘉信 馬場 Microchip electrophoresis apparatus and electrophoresis method using the same
US20030062833A1 (en) * 2001-10-03 2003-04-03 Wen-Yen Tai Mini-type decorative bulb capable of emitting light through entire circumferential face
US6627076B2 (en) * 2001-10-19 2003-09-30 Sandia National Laboratories Compact microchannel system
US7247274B1 (en) 2001-11-13 2007-07-24 Caliper Technologies Corp. Prevention of precipitate blockage in microfluidic channels
US6889468B2 (en) 2001-12-28 2005-05-10 3M Innovative Properties Company Modular systems and methods for using sample processing devices
US7459127B2 (en) * 2002-02-26 2008-12-02 Siemens Healthcare Diagnostics Inc. Method and apparatus for precise transfer and manipulation of fluids by centrifugal and/or capillary forces
US6958119B2 (en) * 2002-02-26 2005-10-25 Agilent Technologies, Inc. Mobile phase gradient generation microfluidic device
US20040029143A1 (en) * 2002-02-28 2004-02-12 Jeffrey Van Ness Cationic polyelectrolytes in biomolecule purification and analysis
EP2581739B1 (en) 2002-03-05 2015-11-04 Caliper Life Sciences, Inc. Microfluidic separation method with combined pressure and voltage control
US7195986B1 (en) * 2002-03-08 2007-03-27 Caliper Life Sciences, Inc. Microfluidic device with controlled substrate conductivity
EP2666849A3 (en) 2002-04-01 2014-05-28 Fluidigm Corporation Microfluidic particle-analysis systems
CA2480200A1 (en) * 2002-04-02 2003-10-16 Caliper Life Sciences, Inc. Methods and apparatus for separation and isolation of components from a biological sample
US20050026134A1 (en) * 2002-04-10 2005-02-03 Bioprocessors Corp. Systems and methods for control of pH and other reactor environment conditions
US7008979B2 (en) * 2002-04-30 2006-03-07 Hydromer, Inc. Coating composition for multiple hydrophilic applications
US20050106714A1 (en) * 2002-06-05 2005-05-19 Zarur Andrey J. Rotatable reactor systems and methods
US7161356B1 (en) 2002-06-05 2007-01-09 Caliper Life Sciences, Inc. Voltage/current testing equipment for microfluidic devices
US7867193B2 (en) * 2004-01-29 2011-01-11 The Charles Stark Draper Laboratory, Inc. Drug delivery apparatus
US20050238506A1 (en) * 2002-06-21 2005-10-27 The Charles Stark Draper Laboratory, Inc. Electromagnetically-actuated microfluidic flow regulators and related applications
US7381317B2 (en) * 2002-08-12 2008-06-03 Beckman Coulter, Inc. Methods and compositions for capillary electrophoresis (CE)
US7001853B1 (en) * 2002-08-30 2006-02-21 Caliper Life Sciences, Inc. Flow control of photo-polymerizable resin
US7094345B2 (en) * 2002-09-09 2006-08-22 Cytonome, Inc. Implementation of microfluidic components, including molecular fractionation devices, in a microfluidic system
US7455770B2 (en) * 2002-09-09 2008-11-25 Cytonome, Inc. Implementation of microfluidic components in a microfluidic system
US6878271B2 (en) * 2002-09-09 2005-04-12 Cytonome, Inc. Implementation of microfluidic components in a microfluidic system
US20040047767A1 (en) * 2002-09-11 2004-03-11 Richard Bergman Microfluidic channel for band broadening compensation
US7329545B2 (en) 2002-09-24 2008-02-12 Duke University Methods for sampling a liquid flow
US6911132B2 (en) * 2002-09-24 2005-06-28 Duke University Apparatus for manipulating droplets by electrowetting-based techniques
US7143785B2 (en) 2002-09-25 2006-12-05 California Institute Of Technology Microfluidic large scale integration
WO2004040001A2 (en) 2002-10-02 2004-05-13 California Institute Of Technology Microfluidic nucleic acid analysis
US7125711B2 (en) 2002-12-19 2006-10-24 Bayer Healthcare Llc Method and apparatus for splitting of specimens into multiple channels of a microfluidic device
US7094354B2 (en) 2002-12-19 2006-08-22 Bayer Healthcare Llc Method and apparatus for separation of particles in a microfluidic device
KR100486730B1 (en) * 2003-01-21 2005-05-03 삼성전자주식회사 A method for measuring a zeta potential by using a alternative potential and a T channel
EP2363025A1 (en) 2003-03-27 2011-09-07 PTC Therapeutics, Inc. Targeting enzymes of the TRNA splicing pathway for identification of anti-fungal and/or anti-proliferative molecules
US6835773B2 (en) * 2003-04-10 2004-12-28 Agilent Technologies, Inc. Use of N-methylurea for electrophoresis of small proteins
ES2317016T3 (en) * 2003-04-14 2009-04-16 Caliper Life Sciences, Inc. REDUCTION OF INTERFERENCE IN A MIGRATION DISPLACEMENT TEST.
AU2004236740A1 (en) * 2003-05-02 2004-11-18 Sigma-Aldrich Co. Solid phase cell lysis and capture platform
US7435381B2 (en) * 2003-05-29 2008-10-14 Siemens Healthcare Diagnostics Inc. Packaging of microfluidic devices
EP1628748A2 (en) * 2003-06-05 2006-03-01 Bioprocessors Corporation Reactor with memory component
US20080257754A1 (en) * 2003-06-27 2008-10-23 Pugia Michael J Method and apparatus for entry of specimens into a microfluidic device
US20040265172A1 (en) * 2003-06-27 2004-12-30 Pugia Michael J. Method and apparatus for entry and storage of specimens into a microfluidic device
JP4996248B2 (en) 2003-07-31 2012-08-08 ハンディーラブ インコーポレイテッド Processing of particle-containing samples
US7347617B2 (en) 2003-08-19 2008-03-25 Siemens Healthcare Diagnostics Inc. Mixing in microfluidic devices
US8095197B2 (en) * 2003-11-03 2012-01-10 Microchips, Inc. Medical device for sensing glucose
JP3828886B2 (en) * 2003-11-25 2006-10-04 アイダエンジニアリング株式会社 Selective surface modification / cleaning method
US7867194B2 (en) 2004-01-29 2011-01-11 The Charles Stark Draper Laboratory, Inc. Drug delivery apparatus
AU2005241080B2 (en) 2004-05-03 2011-08-11 Handylab, Inc. Processing polynucleotide-containing samples
US8852862B2 (en) 2004-05-03 2014-10-07 Handylab, Inc. Method for processing polynucleotide-containing samples
AU2005327211A1 (en) * 2004-06-01 2006-08-17 Microchips, Inc. Devices and methods for measuring and enhancing drug or analyte transport to/from medical implant
EP1765487A1 (en) * 2004-06-07 2007-03-28 Bioprocessors Corporation Reactor mixing
EP1761331A2 (en) * 2004-06-07 2007-03-14 Bioprocessors Corporation Control of reactor environmental conditions
EP1758674A2 (en) * 2004-06-07 2007-03-07 Bioprocessors Corporation Creation of shear in a reactor
US20070036690A1 (en) * 2004-06-07 2007-02-15 Bioprocessors Corp. Inlet channel volume in a reactor
US7211184B2 (en) * 2004-08-04 2007-05-01 Ast Management Inc. Capillary electrophoresis devices
WO2006026248A1 (en) * 2004-08-25 2006-03-09 Sigma-Aldrich Co. Compositions and methods employing zwitterionic detergent combinations
US7497937B2 (en) * 2004-09-03 2009-03-03 Combisep, Inc. Microfabricated chip and method of use
US7413846B2 (en) * 2004-11-15 2008-08-19 Microchips, Inc. Fabrication methods and structures for micro-reservoir devices
CA2595457A1 (en) * 2005-01-25 2006-08-03 Microchips, Inc. Control of drug release by transient modification of local microenvironments
PL1859330T3 (en) 2005-01-28 2013-01-31 Univ Duke Apparatuses and methods for manipulating droplets on a printed circuit board
US8730031B2 (en) 2005-04-28 2014-05-20 Proteus Digital Health, Inc. Communication system using an implantable device
US8802183B2 (en) 2005-04-28 2014-08-12 Proteus Digital Health, Inc. Communication system with enhanced partial power source and method of manufacturing same
US9198608B2 (en) 2005-04-28 2015-12-01 Proteus Digital Health, Inc. Communication system incorporated in a container
EP2392258B1 (en) * 2005-04-28 2014-10-08 Proteus Digital Health, Inc. Pharma-informatics system
US8912908B2 (en) 2005-04-28 2014-12-16 Proteus Digital Health, Inc. Communication system with remote activation
US8836513B2 (en) 2006-04-28 2014-09-16 Proteus Digital Health, Inc. Communication system incorporated in an ingestible product
US7691263B1 (en) * 2005-05-20 2010-04-06 Brigham Young University Monolithic column technology for liquid chromatography
US20070014699A1 (en) 2005-06-23 2007-01-18 Beckman Coulter, Inc, Methods and apparatus for improving the sensitivity of capillary zone electrophoresis
US7323660B2 (en) 2005-07-05 2008-01-29 3M Innovative Properties Company Modular sample processing apparatus kits and modules
US7763210B2 (en) 2005-07-05 2010-07-27 3M Innovative Properties Company Compliant microfluidic sample processing disks
US7754474B2 (en) 2005-07-05 2010-07-13 3M Innovative Properties Company Sample processing device compression systems and methods
WO2007028035A2 (en) * 2005-09-01 2007-03-08 Proteus Biomedical, Inc. Implantable zero-wire communications system
US8883490B2 (en) 2006-03-24 2014-11-11 Handylab, Inc. Fluorescence detector for microfluidic diagnostic system
US10900066B2 (en) 2006-03-24 2021-01-26 Handylab, Inc. Microfluidic system for amplifying and detecting polynucleotides in parallel
US11806718B2 (en) 2006-03-24 2023-11-07 Handylab, Inc. Fluorescence detector for microfluidic diagnostic system
US7998708B2 (en) 2006-03-24 2011-08-16 Handylab, Inc. Microfluidic system for amplifying and detecting polynucleotides in parallel
DK2001990T3 (en) 2006-03-24 2016-10-03 Handylab Inc Integrated microfluidic sample processing system and method for its use
US8088616B2 (en) 2006-03-24 2012-01-03 Handylab, Inc. Heater unit for microfluidic diagnostic system
CN105468895A (en) 2006-05-02 2016-04-06 普罗透斯数字保健公司 Patient customized therapeutic regimens
EP2087589B1 (en) * 2006-10-17 2011-11-23 Proteus Biomedical, Inc. Low voltage oscillator for medical devices
KR101611240B1 (en) 2006-10-25 2016-04-11 프로테우스 디지털 헬스, 인코포레이티드 Controlled activation ingestible identifier
US8709787B2 (en) 2006-11-14 2014-04-29 Handylab, Inc. Microfluidic cartridge and method of using same
EP2069004A4 (en) 2006-11-20 2014-07-09 Proteus Digital Health Inc Active signal processing personal health signal receivers
EP2117713B1 (en) 2006-12-22 2019-08-07 DiaSorin S.p.A. Thermal transfer methods for microfluidic systems
CA2673056A1 (en) * 2006-12-22 2008-07-03 3M Innovative Properties Company Enhanced sample processing devices, systems and methods
US9046192B2 (en) * 2007-01-31 2015-06-02 The Charles Stark Draper Laboratory, Inc. Membrane-based fluid control in microfluidic devices
CA2676407A1 (en) * 2007-02-01 2008-08-07 Proteus Biomedical, Inc. Ingestible event marker systems
CA2676280C (en) 2007-02-14 2018-05-22 Proteus Biomedical, Inc. In-body power source having high surface area electrode
WO2008112578A1 (en) 2007-03-09 2008-09-18 Proteus Biomedical, Inc. In-body device having a deployable antenna
WO2008112577A1 (en) * 2007-03-09 2008-09-18 Proteus Biomedical, Inc. In-body device having a multi-directional transmitter
US20080227209A1 (en) * 2007-03-14 2008-09-18 David Xing-Fei Deng Methods, Kits And Devices For Analysis Of Lipoprotein(a)
US7799656B2 (en) 2007-03-15 2010-09-21 Dalsa Semiconductor Inc. Microchannels for BioMEMS devices
US8540632B2 (en) 2007-05-24 2013-09-24 Proteus Digital Health, Inc. Low profile antenna for in body device
US7500399B2 (en) * 2007-07-02 2009-03-10 The Hong Kong Polytechnic University Piezoresistive strain gauge using doped polymeric fluid
US8182763B2 (en) 2007-07-13 2012-05-22 Handylab, Inc. Rack for sample tubes and reagent holders
US8105783B2 (en) 2007-07-13 2012-01-31 Handylab, Inc. Microfluidic cartridge
US8133671B2 (en) 2007-07-13 2012-03-13 Handylab, Inc. Integrated apparatus for performing nucleic acid extraction and diagnostic testing on multiple biological samples
US9186677B2 (en) 2007-07-13 2015-11-17 Handylab, Inc. Integrated apparatus for performing nucleic acid extraction and diagnostic testing on multiple biological samples
US9618139B2 (en) 2007-07-13 2017-04-11 Handylab, Inc. Integrated heater and magnetic separator
USD621060S1 (en) 2008-07-14 2010-08-03 Handylab, Inc. Microfluidic cartridge
US8324372B2 (en) 2007-07-13 2012-12-04 Handylab, Inc. Polynucleotide capture materials, and methods of using same
US20090136385A1 (en) 2007-07-13 2009-05-28 Handylab, Inc. Reagent Tube
US8287820B2 (en) 2007-07-13 2012-10-16 Handylab, Inc. Automated pipetting apparatus having a combined liquid pump and pipette head system
US8016260B2 (en) 2007-07-19 2011-09-13 Formulatrix, Inc. Metering assembly and method of dispensing fluid
WO2009021233A2 (en) * 2007-08-09 2009-02-12 Advanced Liquid Logic, Inc. Pcb droplet actuator fabrication
FI2192946T3 (en) 2007-09-25 2022-11-30 In-body device with virtual dipole signal amplification
ES2661739T3 (en) 2007-11-27 2018-04-03 Proteus Digital Health, Inc. Transcorporeal communication systems that employ communication channels
CA2717862C (en) 2008-03-05 2016-11-22 Proteus Biomedical, Inc. Multi-mode communication ingestible event markers and systems, and methods of using the same
ES2696984T3 (en) 2008-07-08 2019-01-21 Proteus Digital Health Inc Ingestion event marker data infrastructure
USD618820S1 (en) 2008-07-11 2010-06-29 Handylab, Inc. Reagent holder
USD787087S1 (en) 2008-07-14 2017-05-16 Handylab, Inc. Housing
MY154217A (en) 2008-08-13 2015-05-15 Proteus Digital Health Inc Ingestible circuitry
WO2010036801A2 (en) 2008-09-26 2010-04-01 Michael Appleby Systems, devices, and/or methods for manufacturing castings
US8036748B2 (en) * 2008-11-13 2011-10-11 Proteus Biomedical, Inc. Ingestible therapy activator system and method
SG172077A1 (en) 2008-12-11 2011-07-28 Proteus Biomedical Inc Evaluation of gastrointestinal function using portable electroviscerography systems and methods of using the same
TWI503101B (en) 2008-12-15 2015-10-11 Proteus Digital Health Inc Body-associated receiver and method
US9439566B2 (en) 2008-12-15 2016-09-13 Proteus Digital Health, Inc. Re-wearable wireless device
US9659423B2 (en) 2008-12-15 2017-05-23 Proteus Digital Health, Inc. Personal authentication apparatus system and method
US7927904B2 (en) 2009-01-05 2011-04-19 Dalsa Semiconductor Inc. Method of making BIOMEMS devices
CN102365084B (en) 2009-01-06 2014-04-30 普罗秋斯数字健康公司 Pharmaceutical dosages delivery system
WO2010080843A2 (en) 2009-01-06 2010-07-15 Proteus Biomedical, Inc. Ingestion-related biofeedback and personalized medical therapy method and system
US8100293B2 (en) 2009-01-23 2012-01-24 Formulatrix, Inc. Microfluidic dispensing assembly
EP2362212B1 (en) 2009-03-18 2019-02-20 Agilent Technologies, Inc. PDMA and derivatized cellulose as separation medium for DNA
WO2010111403A2 (en) 2009-03-25 2010-09-30 Proteus Biomedical, Inc. Probablistic pharmacokinetic and pharmacodynamic modeling
MX2011011506A (en) 2009-04-28 2012-05-08 Proteus Biomedical Inc Highly reliable ingestible event markers and methods for using the same.
EP2432458A4 (en) 2009-05-12 2014-02-12 Proteus Digital Health Inc Ingestible event markers comprising an ingestible component
WO2010141131A1 (en) 2009-06-04 2010-12-09 Lockheed Martin Corporation Multiple-sample microfluidic chip for dna analysis
US8558563B2 (en) 2009-08-21 2013-10-15 Proteus Digital Health, Inc. Apparatus and method for measuring biochemical parameters
TWI517050B (en) 2009-11-04 2016-01-11 普羅托斯數位健康公司 System for supply chain management
USD638951S1 (en) 2009-11-13 2011-05-31 3M Innovative Properties Company Sample processing disk cover
USD638550S1 (en) 2009-11-13 2011-05-24 3M Innovative Properties Company Sample processing disk cover
USD667561S1 (en) 2009-11-13 2012-09-18 3M Innovative Properties Company Sample processing disk cover
US8834792B2 (en) 2009-11-13 2014-09-16 3M Innovative Properties Company Systems for processing sample processing devices
UA109424C2 (en) 2009-12-02 2015-08-25 PHARMACEUTICAL PRODUCT, PHARMACEUTICAL TABLE WITH ELECTRONIC MARKER AND METHOD OF MANUFACTURING PHARMACEUTICAL TABLETS
BR112012019212A2 (en) 2010-02-01 2017-06-13 Proteus Digital Health Inc data collection system
KR20170121299A (en) 2010-04-07 2017-11-01 프로테우스 디지털 헬스, 인코포레이티드 Miniature ingestible device
TWI557672B (en) 2010-05-19 2016-11-11 波提亞斯數位康健公司 Computer system and computer-implemented method to track medication from manufacturer to a patient, apparatus and method for confirming delivery of medication to a patient, patient interface device
JP2012029676A (en) * 2010-07-07 2012-02-16 Sony Corp Cartridge for concentrating and collecting nucleic acid, method for concentrating and collecting nucleic acid and method for fabricating the cartridge
AU2011315951B2 (en) 2010-10-15 2015-03-19 Lockheed Martin Corporation Micro fluidic optic design
JP2014504902A (en) 2010-11-22 2014-02-27 プロテウス デジタル ヘルス, インコーポレイテッド Ingestible device with medicinal product
WO2012106501A1 (en) 2011-02-02 2012-08-09 The Charles Stark Draper Laboratory, Inc. Drug delivery apparatus
EP2683291B1 (en) 2011-03-11 2019-07-31 Proteus Digital Health, Inc. Wearable personal body associated device with various physical configurations
BR112013026451B1 (en) 2011-04-15 2021-02-09 Becton, Dickinson And Company system and method to perform molecular diagnostic tests on several samples in parallel and simultaneously amplification in real time in plurality of amplification reaction chambers
ES2870874T3 (en) 2011-05-18 2021-10-27 Diasorin S P A Systems and methods for detecting the presence of a selected volume of material in a sample processing device
WO2012158988A1 (en) 2011-05-18 2012-11-22 3M Innovative Properties Company Systems and methods for valving on a sample processing device
WO2012158990A1 (en) 2011-05-18 2012-11-22 3M Innovative Properties Company Systems and methods for volumetric metering on a sample processing device
WO2015112603A1 (en) 2014-01-21 2015-07-30 Proteus Digital Health, Inc. Masticable ingestible product and communication system therefor
US9756874B2 (en) 2011-07-11 2017-09-12 Proteus Digital Health, Inc. Masticable ingestible product and communication system therefor
CN103827914A (en) 2011-07-21 2014-05-28 普罗秋斯数字健康公司 Mobile communication device, system, and method
USD692162S1 (en) 2011-09-30 2013-10-22 Becton, Dickinson And Company Single piece reagent holder
WO2013049706A1 (en) 2011-09-30 2013-04-04 Becton, Dickinson And Company Unitized reagent strip
WO2013067202A1 (en) 2011-11-04 2013-05-10 Handylab, Inc. Polynucleotide sample preparation device
US9235683B2 (en) 2011-11-09 2016-01-12 Proteus Digital Health, Inc. Apparatus, system, and method for managing adherence to a regimen
US8813824B2 (en) 2011-12-06 2014-08-26 Mikro Systems, Inc. Systems, devices, and/or methods for producing holes
CN104204812B (en) 2012-02-03 2018-01-05 贝克顿·迪金森公司 The external file that compatibility determines between distributing and test for molecule diagnostic test
US9322054B2 (en) 2012-02-22 2016-04-26 Lockheed Martin Corporation Microfluidic cartridge
JP2015534539A (en) 2012-07-23 2015-12-03 プロテウス デジタル ヘルス, インコーポレイテッド Technique for producing an ingestible event marker with an ingestible component
US9268909B2 (en) 2012-10-18 2016-02-23 Proteus Digital Health, Inc. Apparatus, system, and method to adaptively optimize power dissipation and broadcast power in a power source for a communication device
WO2014120669A1 (en) 2013-01-29 2014-08-07 Proteus Digital Health, Inc. Highly-swellable polymeric films and compositions comprising the same
WO2014151929A1 (en) 2013-03-15 2014-09-25 Proteus Digital Health, Inc. Personal authentication apparatus system and method
US10175376B2 (en) 2013-03-15 2019-01-08 Proteus Digital Health, Inc. Metal detector apparatus, system, and method
JP6511439B2 (en) 2013-06-04 2019-05-15 プロテウス デジタル ヘルス, インコーポレイテッド Systems, devices, and methods for data collection and outcome assessment
US9796576B2 (en) 2013-08-30 2017-10-24 Proteus Digital Health, Inc. Container with electronically controlled interlock
EP3047618B1 (en) 2013-09-20 2023-11-08 Otsuka Pharmaceutical Co., Ltd. Methods, devices and systems for receiving and decoding a signal in the presence of noise using slices and warping
WO2015044722A1 (en) 2013-09-24 2015-04-02 Proteus Digital Health, Inc. Method and apparatus for use with received electromagnetic signal at a frequency not known exactly in advance
US10084880B2 (en) 2013-11-04 2018-09-25 Proteus Digital Health, Inc. Social media networking based on physiologic information
EP3077037B1 (en) 2013-12-04 2021-11-17 ReShape Lifesciences Inc. System for locating intragastric devices
US10421072B2 (en) 2014-01-21 2019-09-24 The Board Of Trustees Of The University Of Illinois Wettability patterned substrates for pumpless liquid transport and drainage
US9895248B2 (en) 2014-10-09 2018-02-20 Obalon Therapeutics, Inc. Ultrasonic systems and methods for locating and/or characterizing intragastric devices
US11051543B2 (en) 2015-07-21 2021-07-06 Otsuka Pharmaceutical Co. Ltd. Alginate on adhesive bilayer laminate film
US10350100B2 (en) 2016-04-12 2019-07-16 Obalon Therapeutics, Inc. System for detecting an intragastric balloon
CN109843149B (en) 2016-07-22 2020-07-07 普罗秋斯数字健康公司 Electromagnetic sensing and detection of ingestible event markers
CN109963499B (en) 2016-10-26 2022-02-25 大冢制药株式会社 Method for manufacturing capsules with ingestible event markers
US11207679B2 (en) * 2018-04-13 2021-12-28 Regents Of The University Of Minnesota DNA extraction device
WO2023164258A1 (en) * 2022-02-28 2023-08-31 Perkinelmer Health Sciences, Inc. Methods for separating and detecting double-stranded and single-stranded ribonucleic acid (rna)

Family Cites Families (30)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
SE328715B (en) * 1967-04-20 1970-09-21 Incentive Res & Dev Ab
US4054432A (en) * 1976-06-11 1977-10-18 Wright State University Polymer lined capillary column and method for producing same
US4390403A (en) * 1981-07-24 1983-06-28 Batchelder J Samuel Method and apparatus for dielectrophoretic manipulation of chemical species
US4509964A (en) * 1984-01-04 1985-04-09 The Foxboro Company Fused silica capillary column
US5171534A (en) * 1984-01-16 1992-12-15 California Institute Of Technology Automated DNA sequencing technique
ATE122918T1 (en) * 1987-03-24 1995-06-15 Univ Northeastern HIGHLY EFFECTIVE SELECTIVE ELECTROKINETIC SEPARATIONS ON THE SURFACE OF MOVING CHARGED COLLOIDAL PARTICLES.
US4908112A (en) * 1988-06-16 1990-03-13 E. I. Du Pont De Nemours & Co. Silicon semiconductor wafer for analyzing micronic biological samples
US5089111A (en) * 1989-01-27 1992-02-18 Bio-Rad Laboratories, Inc. Electrophoretic sieving in gel-free media with dissolved polymers
US5096554A (en) * 1989-08-07 1992-03-17 Applied Biosystems, Inc. Nucleic acid fractionation by counter-migration capillary electrophoresis
US5181999A (en) * 1989-11-06 1993-01-26 Applied Biosystems, Inc. Capillary electrophoresis method with polymer tube coating
US5264101A (en) * 1989-11-06 1993-11-23 Applied Biosystems, Inc. Capillary electrophoresis molecular weight separation of biomolecules using a polymer-containing solution
US5126022A (en) * 1990-02-28 1992-06-30 Soane Tecnologies, Inc. Method and device for moving molecules by the application of a plurality of electrical fields
US5935401A (en) * 1996-09-18 1999-08-10 Aclara Biosciences Surface modified electrophoretic chambers
US5332481A (en) * 1991-01-29 1994-07-26 Beckman Instruments, Inc. Capillary electrophoresis using replaceable gels
US5126021A (en) * 1991-07-17 1992-06-30 Applied Biosystems Inc. Low-viscosity polymer solution for capillary electrophoresis
US5498392A (en) * 1992-05-01 1996-03-12 Trustees Of The University Of Pennsylvania Mesoscale polynucleotide amplification device and method
US5304487A (en) * 1992-05-01 1994-04-19 Trustees Of The University Of Pennsylvania Fluid handling in mesoscale analytical devices
US5637469A (en) * 1992-05-01 1997-06-10 Trustees Of The University Of Pennsylvania Methods and apparatus for the detection of an analyte utilizing mesoscale flow systems
US5374527A (en) * 1993-01-21 1994-12-20 Applied Biosystems, Inc. High resolution DNA sequencing method using low viscosity medium
JP3012003B2 (en) * 1993-10-14 2000-02-21 バイオ−ラッド ラボラトリーズ,インコーポレイティド Suppression of electroosmosis during electrophoresis in gel-free polymer media by using a charged polymer
WO1995016910A1 (en) * 1993-12-17 1995-06-22 Perkin-Elmer Corporation Uncharged polymers for separation of biomolecules by capillary electrophoresis
DE69409626T2 (en) * 1994-01-28 1998-11-05 Hewlett Packard Gmbh Plastic capillary for capillary electrophoresis and process for its manufacture
US6001229A (en) * 1994-08-01 1999-12-14 Lockheed Martin Energy Systems, Inc. Apparatus and method for performing microfluidic manipulations for chemical analysis
US5571410A (en) * 1994-10-19 1996-11-05 Hewlett Packard Company Fully integrated miniaturized planar liquid sample handling and analysis device
US5603351A (en) * 1995-06-07 1997-02-18 David Sarnoff Research Center, Inc. Method and system for inhibiting cross-contamination in fluids of combinatorial chemistry device
US5585069A (en) * 1994-11-10 1996-12-17 David Sarnoff Research Center, Inc. Partitioned microelectronic and fluidic device array for clinical diagnostics and chemical synthesis
US5611903A (en) * 1995-03-22 1997-03-18 Analis S. A. Capillary electrophoresis method using initialized capillary and polyanion-containing buffer and chemical kit therefor
US5856174A (en) * 1995-06-29 1999-01-05 Affymetrix, Inc. Integrated nucleic acid diagnostic device
US5858187A (en) * 1996-09-26 1999-01-12 Lockheed Martin Energy Systems, Inc. Apparatus and method for performing electrodynamic focusing on a microchip
US5948227A (en) * 1997-12-17 1999-09-07 Caliper Technologies Corp. Methods and systems for performing electrophoretic molecular separations

Also Published As

Publication number Publication date
JP2002515586A (en) 2002-05-28
US6440284B1 (en) 2002-08-27
US6042710A (en) 2000-03-28
US7081190B2 (en) 2006-07-25
EP1040343A4 (en) 2006-10-18
AU747232B2 (en) 2002-05-09
EP1040343A1 (en) 2000-10-04
JP4144776B2 (en) 2008-09-03
AU1619899A (en) 1999-07-05
CA2309831A1 (en) 1999-06-24
US5948227A (en) 1999-09-07
WO1999031495A1 (en) 1999-06-24
US20020166768A1 (en) 2002-11-14

Similar Documents

Publication Publication Date Title
CA2309831C (en) Improved methods and systems for performing molecular separations
US6787015B2 (en) Methods for conducting electrophoretic analysis
CA2249886C (en) Acrylic microchannels and their use in electrophoretic applications
US6054034A (en) Acrylic microchannels and their use in electrophoretic applications
Kan et al. DNA sequencing and genotyping in miniaturized electrophoresis systems
US20060210995A1 (en) Nanopore analysis systems and methods of using nanopore devices
EP2172771B1 (en) High Speed, high resolution method for capillary electrophoresis
US20050053944A1 (en) Methods and kit for hybridization anaylsis using peptide nucleic acid probes
US20060207880A1 (en) Microfluidic devices and methods of using microfluidic devices
CA2222628A1 (en) Microelectrophoresis chip for moving and separating nucleic acids and other charged molecules
WO1997012995A9 (en) Methods and kit for hybridization analysis using peptide nucleic acid probes
WO2002042500A2 (en) Apparatus and method for electrophoretic microspot concentration
Chen et al. High-throughput DNA analysis by microchip electrophoresis
US6878254B2 (en) Size separation of analytes using monomeric surfactants
US20040101970A1 (en) Treatment solution minimising adsorption and/or elecroosmosis phenomena
US20100051459A1 (en) Denaturant-Free Electrophoresis of Biological Molecules Under High Temperature Conditions
JP3727031B2 (en) Fluorescence-based electrophoresis system for polynucleotide analysis
US20060102480A1 (en) Apparatus and methods for performing electrophoretic separations of macromolecules
Freitag Capillary Gel Electrophoresis
Kan Thermo-responsive and thermo-gelling polymer networks for DNA sequencing and genotyping by capillary and microchip electrophoresis

Legal Events

Date Code Title Description
EEER Examination request
MKLA Lapsed