CA1081591A - Process for mounting tissue sections with an u. v. light curable mounting medium - Google Patents

Process for mounting tissue sections with an u. v. light curable mounting medium

Info

Publication number
CA1081591A
CA1081591A CA282,736A CA282736A CA1081591A CA 1081591 A CA1081591 A CA 1081591A CA 282736 A CA282736 A CA 282736A CA 1081591 A CA1081591 A CA 1081591A
Authority
CA
Canada
Prior art keywords
specimen
solvent
subsequent
materials
polymerizable
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
Application number
CA282,736A
Other languages
French (fr)
Inventor
Leonard Ornstein
Hazel E. Williams
Julius Intraub
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.)
Bayer Corp
Original Assignee
Technicon Instruments Corp
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 Technicon Instruments Corp filed Critical Technicon Instruments Corp
Application granted granted Critical
Publication of CA1081591A publication Critical patent/CA1081591A/en
Expired legal-status Critical Current

Links

Classifications

    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00Sampling; Preparing specimens for investigation
    • G01N1/28Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/30Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
    • G01N1/31Apparatus therefor
    • G01N1/312Apparatus therefor for samples mounted on planar substrates

Abstract

ABSTRACT OF THE DISCLOSURE
Apparatus and method for permanently and protectively mount-ing "wet" thin sections of biological specimens on microscope slides wherein each specimen is initially bathed in a low-volatile solvent containing a low concentration of first polymerizable material.
Such solvent is allowed to evaporate, whereby the liquid polymeri-zable material permeates and protects the specimen. Following solvent evaporation, a second polymerizable material is layered over the specimen. Preferably, the first and second polymeriable materials comprise a mixture of low-volatility, low-viscosity, li-quid acrylic reactomers and a U.V. light-sensitive catalyst system conventional cover slip or other planar transparent member can be positioned over the polymerizable materials, and the same are polymerized by exposure to U.V. radiation. The first and second now-polymerized materials encapsulate the specimen on the microscope slide, become integral and completely hardened and are fully devoid of any solvent. The microscope slide is immediately available for examination.

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Description

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BACKGROUND OF THE INVENTION
1. Field of the Invention This invention relates -to a method and apparatus for providing permanent, optically clear preparations of biological specimens on microscope slides for cytologic, histologic and pathologic study.
2. Description of the Prior Art It is common practice in the medical field to provide thin sections of biological specimens on glass slides for micro-scopic study, such sections have been stained ~_o enhance def-inition for examination), dehydrated and subjected to a mount-ing process. Generally, such mounting process encloses each individual specimen in a medium supported on a microscope slide and wherein it is preserved indefinitely. The present trend is to use a solvent solution of hard synthetic resins as the mounting medium, or mountant. Such resins are chosen so that they (1) do not cause the stains with which the specimens are treated to fade; (2) yellow minimally with age, and ~3) have refractive indices after drying which closely approximate the average refractive index of tissues ~1.530-1.570). If a mountant having a refractive index close to that of the specimen is used, an almost perfect transparency can be achieved. A ~-pinene resin is often used as moun~ant. For a listing of other avail-able mountants, reference is made to table 5-1 on page 98 of Histopathic Technique and Practical Histo-chemistry, by R.D.
Lillie, McGraw-Hill Book Company, New York (1965).
Generally, the mountant is dissolved in a quantity of solvent, to provide proper viscosity for the convenient dispensing of a controlled volume thereof over the specimen. To provide a planar upper surface, necessary for proper microscopic examina-:: ,
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tion, it is common practice to apply a glass cover slip over the dispensed moun-~ant while still in liquid form. To remove all solvent, the presence of which can interfere with the ex-amination process, the slides are subjected to an extended drying process, which may require several days or more. During such process, the slides are placed in a horizontal position, and sometimes within a warm environment, to facilitate sol-vent evaporation. Such solvent evaporation is essential to ~1) harden the mountant, (2) seal the cover slip, so as to per-mit handling of the slides, and (3) avoid optical interferenceby the low refractive index of residual solvent during examina-tion.
At the present time, the mounting and drying processes are done manually in the laboratory. Commonly, after removing the slide from the so-called clearing solution and quickly draining the same, one to several drops of mountant are deposited by the technician over the specimen. The quantity of such mountant should be sufficient to just occupy the space between the cover slip, when'applied, and the specimen.
Immediately, the cover slip is carefully positioned by the technician by being lowered gently upon the mountant. The technique contemplates that one side of the cover slip is placed on the mountant, which is then released to push before it a portion of the mountant as it descends into proper position. Often, the cover slip requires some adjustment by the technician, so as to achieve a thin even film of mountant, to eliminate any entrapped air bubbles from beneath the ` cover slip, and to properly center the cover slip over the -specimen. Any surplus of mountant expressed beyond the-~
cover slip should be quickly and carefully removed, for ph: b~

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example, with absorbent paper, by the techni ~ n.
The most common difficulties encountered in the mounting of specimens are (1) a delayed application of the mountant after the specimen is removed from the clearing agent allows the drying of the specimen, whereby some deterioration of the tissue structure results, (2) the amount of mountant is inappropriate, whereby the resultant film beneath the cover slip is too thick or too thin or the plane of the cover slip is not parallel with that of the microscope slide, and (3) too rapid or careless application of the cover slip traps air bubbles, which become increasingly more difficult to release as the mountant begins to dry.
The prior art procedures, which have been only briefly ;
described, have well-reco~nizedi disadvantages, among which are:
1. Since the specimen can be damaged if allowed to dry, the mountant must be carefully and quickly applied by the technician, 2. Since the solvent in the mountant has a low index of refraction and diffuses only slowly out from under the cover slip during the drying process, the mountant is generally prepared with a minimum of solvent, usually sufficient only to allow convenient dispensing. However, if the placement of the cover slip is delayed, the surface of the mountant can begin to dry, which tends to increase the probability of trapping air bubbles under the cover slip. Accordingly, the cover slip must be both carefully and quickly positioned on the mountant.
3. If an excess of mountant is dispensed, such excess-ph~
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must be e~pressed beyond the edges of the cover slip.
Adequate removal of such excess rnay re~uire the use of an appropriate solvent, which might flow under the cover slip and remove underlying mountant, ulti~lately leaving air spaces which would interfere with the examination.
4. During the drying process, the slide must be set aside on a flat horizontal surface i~or a considerable period. Tilting of the slide during such process often causes the cover slip to slide off the tissue section. Also, since the edges of the mountant extend slightly beyond the periphery of the cover slip and diffusion of solvent from under such cover slip keeps such edges sticky, the slides cannot be stacked for efficient storage until the drying process is complete.
5. Even after a prolonged drying process, a minimum quantity of solvent remains under the central portion of the cover slip. Inasmuch as the solvent has a relatively low refractive index, the optical properties of the slide remain sub-optimal for a long time period.
6. If the tissue specimen should have a high point, a thick layer of mounting medium would be present under the cover slip. Accordingly, the drying process would be delayed and considerable shrinkage of the mounting medium would result as the solvent evaporates.
` Such shrinkage often occurs unevenly and locally, such as to create an air space under the cover slip which would obscure tissue detail.

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OBJECTS OF THE INVENTION
An object of the presen-t invention is to provlde im-proved method and apparatus for mounting a tissue specimen, where-by the total processiny time is very signi~icantly reduced.
An additional object of this invention is to provide improved metllod and apparatus for mounting tissue specimens, whereby the prolonged drying process of prior art procedures is substantially eliminated, such that mechanically stable and dry mounted specimens are immediately available for examination.
Another object of this invention is to provide apparatus and method for immediately achieving optically clear and stable preparations of such specimens having indices o refraction closely approximating that of the specimen.
SUMMARY OF THE INVENTION
According to the present invention, a mountant comprising a mixture of low-volatility, low-viscosity, liquid acrylic ;
reactomers, and an ultra violet~light sensitive catalyst system is employed as the final encapsulation medium. Such mountant remains free-flowing until polymerized, whereupon it becomes a hardened, optically clear, solvent-impermeable and solvent-proof medium fully encapsulating the specimen. Such mountant does not cause the stains to fade and remains colorless and clear. The acrylic reactomers are particularly chosen to provide a final re-fractive index, after polymerization, between 1.530-1.570 (typically 1.550) to match that of the specimen. As is known, refractive index can be controlled, for example, by mixing two components having lower and higher refractive indices re-spectively, and varying proportions, to achieve any intermediate refractive index.
As the dehydrated section must not be permitted to dry, ph: b~
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the specimen-carrying slide is initially t~pically bathed in a low-viscosit~v, high volatility solvent solution of a low concentration of mountant. Such mountant may be either of the same material as the final encapsulation medium or an e~uivalent compatible ma-terial, to yield a same refractive index after polymerization.
The slide is then drained. In contrast to standard or classical procedures, the solvent is allowed to completely evaporate, such that a very thin protective layer of solvent-free but liquid mountant remains over the specimen at this time to prevent drying. As the components of such mountant have an extremely low-volatility, they do not evaporate significantly. Rather, the tissue remains wet and permeated with the mountant. When the solvent has been evaporated, usually within one minute, an appropriate final volume, or layer, of the final encapsula-; ting mountant is placed over the "wet" section and the cove~ slip is leisurely applied. The mountant is then exposed through the cover slip (or microscope slide) to U.V. radiation, typically ~` from a low-wattage flourescent black-light lamp, to effect the polymerization thereof within one minute. Accordingly, successively applied layers of the mountant under the cover slip become integral and completely hardened. Also, the resultant mountant is fully devoid of any solvent which might interfere with the examination process, which can be immediately e~fected.
Also, an advantage of the present invention is that the small excess of mountant expressed from beneath the cover slip re~
mains liquid, since it remains exposed to oxygen in the atmosphere, `
which acts as a polymerization inhibitor to thin layers of reactomers, as is well known. Accordingly, the microscope slide can be thoroughly washed with an appropriate solvent to re-move excess liquid mountant expressed beyond the cover slip ph: b~ r~

without affecting the polymerized mountant or can be wiped dr~ .
by the technician, so as to be immediately ready for examination ;
or stacked for storage.
The present invention, therefore, generally contemplatesthe dual application of mountant over the specimen. The first 1 0 ' ` `''i :
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application of mountant is effected with a highly volatile solvent solution of a low concentration of mountant immediately following removal of the slide from the clearing solvent and the second application is effected with pure mountant. Each such mountant is polymerizable, preferably sensitive to U.V.
light, and compatible with the other. Regarding the first application, the solvent is allowed to evaporate completely from the uncovered mountant, whereby the specimen is fully protected from drying and permeated by the mountant, which is still in liquid form. During the second application, any additional necessary volume of mountant is applied over the first layer of mountant, followed by a cover slip, and polymerization of both layers effected concurrentIy. Accordingly, ;~
the individually applied layers of mountant become integral ~ and are fully devoid of all solvent, so as to be capable after - polymerization of being handled immediately for examination.
In one particular aspect the present invention provides `
a method of mounting a stained thin section of a biological specimen, comprising the steps of providing a thin layer of ~`~
polymerizable material of low volatility over the surface of a solvent-wet specimen supported on a transparent substrate, so as to permeate said specimen, substantially totally evaporating the solvent in said specimen, covering said specimen subsequent to the evaporation of said solvent with a planar member which is oxygen-impermeable, and polymerizing said thin layer of materiàl by exposure to U.V. radiation, said thin layer of material preventing damage to said specimen resulting from drying thereof prior to covering with said planar member, said thin layer of material further being transparent when polymeriæed.
In another particular aspect the present invention provides a method of mounting a stained thin section of biological specimen, comprising the steps of bathing a biological specimen, g_ -previously wet with a first volatile solvent and positioned on one s~rface of a transparent substrate, with a second volatile solvent containing a low concentration of a first polymerizable material of low volatility to permeate said specimen, sub-stantially totally evaporating any of said first and secona solvents, providing a thin layer of a second polymerizable material over said specimen and said first material and subsequent to the substantially total evaporation of said first and second solvents, covering said specimen and said layer of second material with a planar member which is oxygen-impermeable, said first material preventing damage to said specimen resulting from drying thereof prior to covering with said planar member, and polymerizing said first and second materials by exposure to U.V. radiation, said first and second materials further being transparent when polymerized.
In a further particular aspect the present invention provides a system for mounting a thin section of a biological ;
: specimen which has been previously wetted by a volatile solvent and is supported upon a transparent carrier, said system comprising:
means for advancing said supported solvent-wet specimen along a feed path past a number of processing stations;
a first processing station disposed along said feed path for applying a polymerizaable material over said solvent-wet specimen before said solvent of said solvent-wet specimen has evaporated;
. a second processing station disposed along said feed path for covering said material-covered specimen with an optically smooth planar member which is oxygen-impermeable, ~ .
subsequent to the total evaporation of said solvent ~rom said solvent-wet specimen; and a third processing station disposed along said feed path for polymerizing, by exposure to ultraviolet light, said polymerizable material subsequent to covering said 1/ ~ ~ -9a-material-covered specimen with said planar member, said polymerizable material being transparent when polymerized.
In yet a further particular aspect the present invention provides a system for mounting a thin section of a biological specimen which has been previously wetted by a first volatile solvent and is supported upon a transparent carrier, said syste~ comprising:
a pretreat processing station for applying a first polymerizable material to said specimen by bathing said r previously wetted specimen with a second volatile solvent " containing a low concentration of a first polymerizable material of low volatility to permeate said specimen;
means for advancing said supported solvent-wet specimen along a feed path past a number of subsequent processing `~
stations;
a first processing station disposed along said feed `
path for applying a second polymerizable material over said , specimen and said first polymerizable material; ;
a second processing station disposed along said feed `~
path for covering said material-covered specimen prior to evaporation of said first and second solvents with an optically smooth planar member which is oxygen-impermeable; and a third processing station disposed along said feed path for polymerizing, by exposure to ultraviolet light, both said first and second polymerizable materials subsequent to covering said material-covered specimen with said planar ;~
member, said first and second polymerizable materials being transparent when polymerized.
BRIEF DESCRIPTION OF THE DRAWINGS
Fig. lA is an isometric view of the apparatus for mounting tissue specimens on microscope slides and embodying the invention;

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Fig. ls is a sectional view of ancillary apparatus to be used in conjunction with the apparatus of Fig. lA.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT
Referring now to the drawings, an apparatus is illustrated in Fig. lA, which embodies the method of the present invention.
Such apparatus comprises a feeding arrangement 1 for successively delivering microscope slides 3, each supporting a tissue specimen 5, which is to be mounted for examination. Each microscope slide 3 includes a label portion 7, identifying the corresponding tissue specimen 5. Initially, each tissue - specimen 5 has been positioned on a corresponding microscope slide 3 by conventional ~' .

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techniques, dried, deparaffined, hydra-ted, stained, dehydrated, cleared and then bathed in a li~uid comprising a low concentra-tion (1~ to 25%) solution of mountant in a lo~-viscosity, hiyh-volatility solvent. As illustrated in Fig. lB, such liquid may be contained within a container 9, in-to which each microscope slide 3 would be immersed by the technician, immediately following ; the conventional clearing step, whereby the specimen is left wet with a volatile solvent, e.g., xylene. The liquid in con-tainer 9 might consist, for example, of xylene, toluene, or Freon~
TF, and an appropriate mountant. Bathing in container 9 provides, after removal, a thin layer of liquid over tissue specimen 5.
The solvent in such liquid is allowed to completely evaporate, such that a very thin protective layer of solvent-free liquid moun-tant remains over both surfaces of the microscope slide. At this time, the tissue specimen 5 remains wetted and permeated through-out with the mountant. Each microscope slide 3 is positioned, in turn, on the feeding arrangement 1, so as to be passed successively to a layering unit 11.
- Layering unit 11 functions to provide a second layer of the same mountant over tissue specimen 5, which mountant comprises a mixture of low-volatility, low-viscosity liquid acrylic reactomers such as monomers or oligomers of methacrylates, acrylates and vinyls and, also a catalyst sensitive to U.V. light, such as benzoin, benzoin ethers, acetophenones, or Michler's Ketone.
Additionally, the layering unit 11 functions to expose the layers of mountant to U.V. radiation peaked near the absorption maximum of the catalyst, whereby both layers are poly~Lerized in on-line fashion. Subsequently, microscope slides 3 are passed, in turn, through a wash station 13, wherein the surfaces of each slide are ph ~v ~ 15~
washed to remove all portions of unpolymerized mountant remaining thereon. The washed microscope slides 3 are subsequently passed through a drying station 15 to accelerate evaporation of the washing liquid and subsequently to a stacker arrangement 17 for accumulating the fully-processed r.icroscope slides 3, which are inunediately available for examination. The total processing time required for completely mounting, washing and drying each microscope slide from the time introduced into layering unit 11 is approximately two minutes.
To more particularly describe the apparatus, each micro-scope slide 3 supporting a tissue specimen 5 and after manual immersion into and removal from container 9 is manually positioned by the technican onto feeding arrangement l. While the bathing and positioning of the microscope slides has been described as ~`
effected manually, the present invention contemplates that structure for effecting the same in on~line fashion could be included as an integral part of the feeding arrangement 1.
Feeding arrangement l, only a portion of which is illustrated in Fig. lA., comprises a parallel belt assembly comprising endless belts l9 and 21 and a suitably driven roller 23. Prefer-ably, roller 23 along with the non-illustrated associated idler roller are recessed along the central surface portions to define shoulders upon which belts l9 and 21, respectively, are trained.
Each microscope slide 3 is positioned on parallel belts l9 and 21, such that the tissue specimen 5 is located on the lower surface and disposed between such belts. Additionally, a pair of parallel guides 55, only one of which is shown, are positioned in near adjacency to belts 19 and 21 to ensure the proper position-ing and orientation of each slide 3 thereon.
The passage of each microscope slide 3 along feeding ~h . L ._ arrangement 1 is sensed by means of~a feeIer arrangement which initiates operation of the apparatus with respect to processing of each such slide. Such feeler arrangement comprises a finger 25 mounted at one end of the rotating shaft 27. Shaft 27 is supported by bearings 28, such that finger 25 extends downward through the spacing between belts 19 and 21. The passage of a microscope slide 3 forces finger 25 to rotate shaft 27 which, in turn, operates microswitch 29.
Microswitch 29, when operated, initiates operation of the layering unit 11, as now described. Layering unit 11 comprises a horizontal platen, or support, member 31 disposed bet~7een idler rollers 35 and 37 and over which a continuous optically smooth `
ribbon 33 is drawn. Ribbon 33 is supplied from supply spool 39 and advanced by means of a take-up spool 41. Appropriate tension is maintained on ribbon 33 by idler rollers 36 and 38. Operation `-of the take-up spool 41 is initiated by operation of microswitch 29. Additionally, operation of the microswitch 29 initiates dispensing of mountant over the upper surface 43 of ribbon 33 for ultimate encapsulation of the tissue specimen 5 on the micro-scope slide 3 which is then being advanced to layering unit 11.
A dispenser 45 is positioned over and adjacent to the ;
upper surface 43 of ribbon 33 and connected through a ~alve 47 and along conduit 49 to a pressurized source 51 of mountant. Valve 47 is operated by the closure of microswitch 29, with appropriate delay, to dispense a continuous bead 53 of mountant onto surface 43 of ribbon 33. The length of bead 53 should preferably be ;
as long as the length of a microscope slide. Also, the mountant is dispensed at a rate to ultimately form a thin layer thereof between the opposing surfaces of the microscope slide 3 and ribbon 33, as hereinafter described.

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~o~S~l As illustrated, each microscope sli~e 3, in turn, is passed from over the belts 19 and 21 of feeding arrangement 1 onto a first ramp arrangement comprising guides 55, only one o~
which is illustrated. Guides 55 are formed in mirrored L~shaped fashion to define a shoulder portion, for supporting microscope slide 3 and a guide portion for orienting such slide during passage. The inclination of guides 55 is such as to gravitational-ly feed each microscope slide 3 onto ribbon 33. The advance of ribbon 33 causes the mlcroscope slide 3 to be layered over the bead S3 of mountant dispensed on the surface o~ such ribbon. As ribbon 33 is initially contactea by the leading end of each micro-scope slide 3, the layering of such slide expresses mountant to the edges and ensures that the same is spread between the entire opposing sur~aces of the slide and ribbon. In this connection, the ribbon 33 functions as would the cover slip in the conventional technique. The volume of mountant dispensed upon ribbon 33 should be only slightly in excess of that required to ensure the forma-tion of a continuous layer of mountant between such opposing sur-faces. The positive spreading of mountant between the opposing surfaces of the microscope slide 3 and ribbon 33 and the planarity of the ribbon ensure that any surface irregularities in the first layer of mountant are filled and that the surface of the resul-ting, or final, layer of mountant is optically smooth and planar.
As ribbon 33 is further advanced by the action of take-up spool 41, the microscope slide 3 is carried through a chamber 57, which includes an appropriate black-light lamp, which is not illustrated, for polymerizing the mountant layers between opposing surfaces of the microscope slide 3 and ribbon 33. Such lamp may be energized by microswitch 29 concurrently with the energization of take-up spool 41. In chamber 57, the mountant ph b~
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layers disposed between ribbon 33 and slide 3 are fully poly-. ;
- merized.
As illustrated, the direction of ribbon 33 after pas-sing over roller 37 is reversed, because of the location of tension roller 38, whereby the ribbon is peeled from the surface of the now-polymerized mountant and gathered on the take-up spool 41. The now-exposed surface of the mountant encapsulating tissue specimen 5 is essentially optically smooth and planar, since defined by ribbon 33. However, any slight excess of mountant -expressed from between ribbon 33 and microscope slide 3 as well as any of the protective layer carried on the surfaces of the :~
slide are unreacted, inasmuch as polymerization thereof has been inhibited by exposure to oxygen in the atmosphere. Subsequently, each microscope slide 3 is passed onto a second ramp arrangement as defined by guides 55. The microscope slide 3 is fed gravi-tationally onto a moving parallel belt assembly comprising belts 61 and 63. Belts 61 and 63 are trained over shoulders ,.
defined at opposite ends of a suitably driven roller 65 and a corresponding idler roller 67, the central surface portions of each being preferably recessed. Preferably, the driven roller 65 is actuated by microswitch 29, with suitable delay, concurrently with the delivery of a microscope slide 3.
Wash station 13 includes a chamber 69 compxising openings disposed on opposite wall portions 71 and 73 for accomo-dating the endless belts 61 and 63. Portions 75 of guides 55 maintain the orientation of the microscope slide 3 on belts 61 and 63, during passage through chamber 69 on belts 61 and 63.
An appropriate washing fluid, for example, Freon TF, for removing unreacted mountant from the surfaces of microscope :
slide 3 is supplied from container 77. Although not illustrated, ph~

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container 77 also includes a positive dlsplacemen~ pump, which is energized by microswitch 29 concurrently ~Jith driven roller 65. The outlet of such pump is connected to conduit 79, which is connected to two branch conduits 81 and 83. The outlets of branch conduits 81 and 83 are arranged to direc-t the pumped wash fluid over the lower and upper surfaces, respectively, of each microscope slide 3 as it is passed through chamber 69.
Preferably, the outlets of branch conduits ~1 and 83, respec-tively, are structured to direct a spray substantially normal to the lower and upper surfaces of the microscope slide 3 be-ing passed therethrough. The wash liquid is subsequently collec-ted at the bottom of chamber 69 and passed along drain 85 and conduit 87 to container 77 for recirculation.
As a microscope slide 3 emerges from chamber 69, it is carried by belts 61 and 63 between blowers 88 and 89, which have been energized concurrently with driven roller 65 and the pump contained within container 77. Blowers 88 and ~9 are disposed to direct warm air over opposite surfaces of the microscope slide 3, to effectively evaporate any residual wash liquid remaining thereon. At this time, the mounting of the tissue specimen 5 on such microscope slide 3 has been completed, and the slide is immediately available for examination.
Subsequent to the drying process, the microscope slide 3 is passed from belts 61 and 63 into a stacking arrangement 17.
Stacking arrangement 17 comprises a bin 91 provided with an integral ramp 93 for receiving microscope slides 3 passed, in turn, from belts 61 and 63. The end of guides 55 cooperate with ramp 93 in maintaining proper orientation of such slides.
Bin 91 is provided a false bottom 97 supported on a spring 99, ph: b r~

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- for minimizing the drop of the microscope slides 3 ~irected thereto. Pre~erably, bin 91 is a four-sided structure, two wall sections having been removed to illustrate the stacking - of the aCcumulated microscope slides 3. Inasmuch as the mount-ant encapsulating the tissue specimen 5 on each microscope slide 3 has been fully reacted and dried, such immediate stack-ing of such slides does not affect the optical properties there-of nor cause them to stick together.
As the apparatus of Fig. lA has been described with respect to mounting of tissue specimens on microscope slides on a discrete basis, it is evident that the components, above described which are controlled by microswitch 29 are operative for a time sufficient to process the individual microscope slides, as they are passed through the apparatus. Of course, it would be evident that each of these same components could be operated on a continuous basis for the mounting of tissue specimens which are directed successive and automatically `
along feeding arrangement 11.
While the presentl~ preferred embodiment of the invention has been illustrated and described, it is apparent to those skilled in the art that the apparatus and method are susceptible to various changes and details without departing from the principles thereof.

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Claims (33)

THE EMBODIMENTS OF THE INVENTION IN WHICH AN EXCLUSIVE PROPERTY
OR PRIVILEGE IS CLAIMED ARE DEFINED AS FOLLOWS:
1. A method of mounting a stained thin section of a biological specimen, comprising the steps of: providing a thin layer of polymerizable material of low volatility over the surface of a solvent-wet specimen supported on a transparent substrate, so as to permeate said specimen, substantially totally evaporating the solvent in said specimen, covering said specimen subsequent to the evaporation of said solvent with a planar member which is oxygen-impermeable, and polymerizing said thin layer of material by exposure to U.V. radiation, said thin layer of material preventing damage to said specimen resulting from drying thereof prior to covering with said planar member, said thin layer of material further being transparent when polymerized.
2. The method of Claim 1, wherein said material exhibits when polymerized a refractive index which approaches that of said specimen.
3. A method of mounting a stained thin section of biological specimen, comprising the steps of: bathing a biological specimen, previously wet with a first volatile solvent and positioned on one surface of a transparent substrate, with a second volatile solvent containing a low concentration of a first polymerizable material of low volatility to permeate said specimen, substantially totally evaporating any of said first and second solvents, providing a thin layer of a second polymerizable material over said specimen and said first material and subsequent to the substantially total evaporation of said first and second solvents, covering said specimen and said layer of second material with a planar member which is oxygen-impermeable, said first material preventing damage to said specimen resulting from drying thereof prior to covering with said planar member, and polymerizing said first and second materials by exposure to U.V. radiation, said first and second materials further being transparent when polymerized.
4. The method of Claim 3 comprising the further step of polymerizing said first and second materials concurrently.
5. The method of Claim 3, wherein at least said second material includes a U.V. light sensitive catalyst system, and comprising the further step of exposing at least said second material to U.V. radiation to effect polymerization thereof.
6. The method of Claim 3, comprising the further step of expressing excess of said first and second materials from between said planar member and said substrate.
7. The method of Claim 3, comprising the further step of polymerizing only portions of said first and second materials located between said substrate and said planar member.
8. The method of Claim 3, comprising the further step of washing said substrate to remove unpolymerized first and second materials from the surface thereof.
9. The method of Claim 3, comprising the further step of removing said planar member subsequent to the polymerization of said first and second materials.
10. The method of Claim 3, wherein said first and second materials each exhibits, when polymerized, a refractive index which approaches that of said specimen.
11. The method of Claim 10, wherein said first and second materials exhibit a same refractive index after polymerization.
12. A system for mounting a thin section of a biological specimen which has been previously wetted by a volatile solvent and is supported upon a transparent carrier, said system comprising:
means for advancing said supported solvent-wet specimen along a feed path past a number of processing stations;
a first processing station disposed along said feed path for applying a polymerizaable material over said solvent-wet specimen before said solvent of said solvent-wet specimen has evaporated;

a second processing station disposed along said feed path for covering said material-covered specimen with an optically smooth planar member which is oxygen-impermeable, subsequent to the total evaporation of said solvent from said solvent-wet specimen; and a third processing station disposed along said feed path for polymerizing, by exposure to ultraviolet light, said polymerizable material subsequent to covering said material-covered specimen with said planar member, said polymerizable material being transparent when polymerized.
13. The system of Claim 12, wherein said thin layer of polymerizable material contains a mixture of low viscosity liquid acrylic reactomers and an ultraviolet light sensitive catalyst system.
14. The system of Claim 12, wherein said thin layer of polymerizable material provides a refractive index when polymerized substantially within the range of 1.530-1.570.
15. The system of Claim 13, wherein the acrylic reactomers are selected from a group consisting of methacrylate, acrylate and vinyl monomers and oligomers.
16. The system of Claim 12, further including means disposed along said feed path for removing said planar member from over said material-covered specimen subsequent to polymerization of said polymerizable material.
17. The system of Claim 12, further including means disposed along said feed path for washing said carrier subsequent to polymerization of any said polymerizable material which was not contained under said oxygen-impermeable planar member, and therefore remained unpolymerized.
18. The system of Claim 17, further including means disposed along said feed path for drying said carrier subsequent to washing thereof.
19. The system of Claim 12, wherein means are provided for sequentially processing, in turn, a plurality of solvent-wet specimens respectively supported upon individual transparent carriers.
20. The system of Claim 19, further including means for accumulating each carrier subsequent to drying thereof.
21. A system for mounting a thin section of a biological specimen which has been previously wetted by a first volatile solvent and is supported upon a transparent carrier, said system comprising:
a pretreat processing station for applying a first polymerizable material to said specimen by bathing said previously wetted specimen with a second volatile solvent containing a low concentration of a first polymerizable material of low volatility to permeate said specimen;
means for advancing said supported solvent-wet specimen along a feed path past a number of subsequent processing stations;
a first processing station disposed along said feed path for applying a second polymerizable material over said specimen and said first polymerizable material;
a second processing station disposed along said feed path for covering said material-covered specimen prior to evaporation of said first and second solvents with an optically smooth planar member which is oxygen-impermeable; and a third processing station disposed along said feed path for polymerizing, by exposure to ultraviolet light, both said first and second polymerizable materials subsequent to covering said material-covered specimen with said planar member, said first and second polymerizable materials being transparent when polymerized.
22. The system of Claim 21, wherein said first and second polymerizable materials contain a mixture of low viscosity liquid acrylic reactomers and an ultraviolet light-sensitive catalyst system.
23. The system of Claim 21 wherein said first and second polymerizable materials provide a refractive index when polymerized substantially within the range of 1.530-1.570.
24. The system of Claim 22, wherein the acrylic reactomers are selected from a group consisting of methacrylate, acrylate and vinyl monomers and oglimers.
25. The system of Claim 21, wherein the means for covering said specimen includes a continuous, movable ribbon.
26. The system of Claim 25, further comprising means for removing said ribbon from a surface of said second polymerizable material following polymerization thereof.
27. The system of Claim 21, further including means for washing the transparent support means subsequent to polymer-ization of said first and second materials.
28. The system of Claim 27, further including means for drying the transparent support means subsequent to washing thereof.
29. The system of Claim 21, further including means disposed along said path for removing said planar member from over said specimen subsequent to polymerization of said polymerizable material.
30. The system of Claim 21, wherein means are provided for sequentially processing, in turn, a plurality of solvent-wet specimens, respectively supported upon individual transparent carriers.
31. The system of Claim 30, further including means disposed along said path for washing each carrier, in turn, subsequent to polymerization of said polymerizable materials.
32. The system of Claim 31, further including means for drying each carrier, in turn, subsequent to washing thereof.
33. The system of Claim 32, further including means for accumulating each carrier, in turn, subsequent to drying thereof.
CA282,736A 1976-12-10 1977-07-14 Process for mounting tissue sections with an u. v. light curable mounting medium Expired CA1081591A (en)

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AU (1) AU501982B2 (en)
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DE2754351C2 (en) 1989-08-31
DE2754351A1 (en) 1978-06-15
FR2373805B1 (en) 1984-02-24
NL7710904A (en) 1978-06-13
JPS5372639A (en) 1978-06-28
FR2373805A1 (en) 1978-07-07
JPS626177B2 (en) 1987-02-09
SE7708010L (en) 1978-06-11
GB1586618A (en) 1981-03-25
US4120991A (en) 1978-10-17
SE439696B (en) 1985-06-24
IT1091166B (en) 1985-06-26
AU2789277A (en) 1979-03-08
AU501982B2 (en) 1979-07-05
BE860067A (en) 1978-04-25
CH626174A5 (en) 1981-10-30

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