Tissue retrieval, storage, and explant culture device for the derivation of stem cells

A device is provided for securing a tissue sample from biological material. The tissue sample is housed in bottom and top platens that are configured to promote fluid communication between the tissue sample and the exterior environment to permit transport or cryogenic fluid to contact the sample. Additionally, diskette assemblies may be provided within the platens that permit sub-samples to be separated without directly handling the tissue sample. The diskette assemblies may also be configured to promote fluid communication with the sub-sample housed therein.

TECHNICAL FIELD

The present disclosure generally relates to methods and apparatus for obtaining tissue samples from biological materials.

BACKGROUND

Within the normal reproductive process, the gestation of a fetus within the female of the species typically occurs within a gestational sac. The gestational sac is comprised of a pair of juxtaposed membranes comprising an outer structure, such as the chorionic membrane (chorion) that forms an outer margin, and an inner structure, such as the amniotic membrane (amnion) which is closest proximity to the fetus. In childbirth following partuation of the gestational sac, commonly referred to as afterbirth, the membrane structure is typically disposed as medical waste. However, medical discoveries are increasingly finding valuable biomedical materials in the tissues of these materials, and specifically the amnion.

The first known clinical uses of the amnion began a century ago for the treatment of various wounds including burn trauma and skin ulcerations on the surface. Extended usage of the intact amnion and chorion tissue came in the 1950's with more focused procedures to treat skin burn sites. The value of amnion was exemplified in the 1960's with a proposal to establish an amnion bank.

Contemporary medicine has seen a relative explosion in the utility of the amnion tissue in the areas of treatment for ocular disorders and thermal and chemical burns. In these treatments, intact amnion tissue is literally transposed directly over the trauma or wound site and has been proven to promote faster healing and alleviate pain. Today the most prevalent use of amnion tissue is in surgical procedures involving the eye.

Recent research is shifting the primary focus of medical attributes to the area of stem cell derivation. The concept of “Regenerative Medicine”, which is replacing, repairing and reconstructing diseased tissue and organs by stem cell therapies, is making rapid clinical progress. Multiple stem cell populations have been identified in the amnion and chorion. When derived at child birth, the benefits can be greater because of their pristine nature due to a lack of exposure to environmental toxins. Any or all derived stem cells may have future clinical utility and may play an extremely large and beneficial role in regenerative medicine.

An alternate and proven method of stem cell derivation in current practice today, and also practiced for the benefit of regenerative medicine, is that of harvesting the placental umbilical cord blood. By this practice, a single sample of blood is harvested from the umbilical cord at birth. The harvested blood is then processed to derive the appropriate beneficial stem cells and the sample is frozen for future use. The ability to cryogenically freeze living cells for thawing and renewal at a later time is a well proven and common practice for many biological materials and is commonly used for procedures such as in vitro fertilization of embryos implanted to induce pregnancy. Cord blood, amnion and chorion tissues are likewise proven biological materials that are conducive to being frozen for an unlimited time. The stem cells derived from cord blood are typically most appropriate for the treatment of hematopoietic (i.e., blood-related) illnesses, and the singular sample retrieved will typically be saved for the future benefit of the donor. By contrast, it has been found that amnion derived stem cells possess qualities that become quite beneficial for treatment of illnesses other than hematopoietic, such as skeletal and cardiovascular. Fundamentally, umbilical cord blood produces hematopoietic stem cells and amnion and chorion produce mesenchymal and epithelial stem cells, all of which are of growing benefit to the healthcare practice of regenerative medicine and are capable of being harvested and preserved for future benefit.

With the growing number of possible clinical applications for amnion and chorion, the devices and methods used to retrieve and preserve these tissues are of increasing interest to the health care community. As new procedures are developed, the devices must be adapted and may advantageously facilitate, optimize, and simplify retrieval and storage of amnion and chorion directly from the afterbirth.

SUMMARY OF THE DISCLOSURE

Devices and methods are disclosed for isolating biological materials and placing them in a position highly conducive to long term storage, thereby preserving the materials intact and preserving them for use by future generations. The benefits of banking and preserving these materials to aid in future medical procedures will only increase in time due to the technological advances in the healthcare industry.

According to some aspects of this disclosure, a device is provided for retrieving, capturing, transporting, and cryogenically storing tissue samples from biological material. In some applications, the device may be used to harvest and store amnion or chorion tissue samples.

Efficient retrieval and handling of amnion and chorion membranes is complicated by the physical characteristics of the membranes. The amnion and chorion membranes are typically extremely thin, possess a level of tensile attributes, but also exhibit a notch sensitive behavior which makes the membranes susceptible to tearing and subsequent curling up upon itself if not properly handled. Tearing and curling of the membranes can destroy cellular viability, and therefore may be deleterious to future clinical value.

Additionally, the amnion and chorion are living membranes when harvested at childbirth, and therefore they should be handled, transported, and preserved correctly to maximize the yield of living cells that are obtained and preserved for future use. Therefore, in addition to folding, or curling back upon itself, or any act whereby the tissue surface is touched by itself, or other capturing elements causing harmful effects, similar damage can be caused by allowing the membrane surface to dry.

In view of the foregoing, according to certain aspects of this disclosure a device may be provided having a working platform upon which to secure the membrane in a way that spreads it out straight, flat and secure. At birth, the amnion and chorion are intimately attached and therefore may require manual separation. In some embodiments, therefore, the methods and devices may secure a leading edge of one membrane (such as the amnion) on a platen while the membranes are separated. After separation, the entire amnion sample may remain on the platen.

The methods and devices disclosed herein may also create multiple smaller sub-samples from a single, larger sample. For example, a first or bottom platen assembly may be capable of being reconfigured into multiple sub-platen assemblies, so that a large sample provided on the bottom platen can be simply and efficiently separated into multiple sub-samples disposed on sub-platen assemblies. Generating multiple sub-samples from a single sample typically requires cutting or tearing of the tissue while simultaneously stabilizing the assembly so that it does not contract or curl upon itself. Accordingly, the methods and devices disclosed herein may include a second or top platen assembly configured to retain the straight and flat nature of each individual sub-sample both prior to and after separation. The top platen assembly is positioned over the bottom platen assembly, and guides are provided to consistently and accurately locate and mates the top and bottom platens with the tissue disposed therebetween, all while facilitating separation into multiple sub-assemblies carrying sub-samples at a later time, such as prior to cryogenic storage.

According to additional aspects of the present disclosure, the methods and devices disclosed herein may further provide a liquid containment of the mated top and bottom platens to preserve the sample as it is transported to a cryogenic storage facility intact.

Therefore, all means of capturing the flat sheet of tissue is accomplished by interrupted and circuitous surfaces that facilitate the flow and intimate contact of a transport liquid with the membrane top and bottom surfaces within a leak proof container device.

According to further aspects of the present disclosure, the methods and devices disclosed herein may enable removal of the mated top and bottom platen assembly from a transport containment, and permit easy separation of the larger tissue sample into multiple, individual, smaller sub-samples. To that end, each sub-sample may be secured in a manner that retains the flatness of the samples. In some embodiments, diskette assemblies may be provided for securing sub-samples, with each diskette assembly including a top diskette and a bottom diskette. Individual top diskettes may be mechanically coupled to the top platen, while individual bottom diskettes may be mechanically coupled to the bottom platen. In order to create multiple individual and smaller samples from a single large sample, the mated platen assembly may be removed from the transport container and placed on a flat surface. The multiple top diskettes, which protrude from the top platen, may be pushed downward and seated into the bottom platen and respective bottom diskettes. When all top diskettes are seated in their respective bottom diskettes, the top platen may be lifted and removed from the assembly. The array of multiple diskette assemblies may remain loosely attached in the exposed bottom platen. Each diskette assembly may encompass an associated portion of the tissue sample, while some or all to the diskette assemblies may continue to be adjoined by connecting membrane of the sample. A gap may be provided between adjacent diskette assemblies that permits a cutting tool, such as a scalpel, to be inserted to cut the connecting membrane. Alternatively, the diskette assemblies may be manually removed from the bottom platen in such a way to tear the connecting membrane and free each individual diskette assembly while the associated sub-sample contained within the separated diskette assembly remains secure and intact.

According to additional aspects of the present disclosure, each diskette may be placed into a vial for cryogenic storage. A cryogenic preservative solution can be added to the vial, while the diskette assemblies limit surface contact with the membrane, retain membrane flatness, and allow maximum solution intimacy with all surfaces for optimal preservation.

In some embodiments, one of the top and bottom diskettes may incorporate a sharp cutting element configured to cut through the sub-sample as the top diskette is mechanically seated on the bottom diskette.

DETAILED DESCRIPTION

The methods and devices disclosed herein enable the successful retrieval of a large tissue sample from biological material in a sterile hospital environment. The tissue sample is captured with limited trauma and degradation to the material while encapsulating it for further transfer and ultimate preservation while in a living state. In particular, the methods and devices disclosed herein may be used to retrieve and preserve amnion tissue that is presented only during the birth of the species. A sample of living amnion tissue is deemed to comprise a highly beneficial population of stem cells, and can therefore be frozen in time to preserve cell viability for later utilization. The methods and devices disclosed herein may also facilitate cryogenic storage and return to maximum living cell viability.

Referring now to the drawings,FIG. 1illustrates an embodiment of a tissue securing device20. The issue securing device20may include a bottom platen22and a top platen24. The bottom and top platens22,24may be provided as separate assemblies so that biological material may be positioned between the platens. Accordingly, when the top platen24is joined to the bottom platen22, a tissue sample26may be secured by the assembly, as described in greater detail below.

The bottom platen22is shown inFIG. 2as having a generally tray-like appearance formed by a bottom platen base30and a bottom platen side wall32. The bottom platen base30defines a bottom platen base inner surface34, while the bottom platen side wall32extends upwardly from a perimeter of the bottom platen base30. Together, the bottom platen side wall32and bottom platen base30define a bottom platen base receptacle36. As best shown inFIG. 3, a flange38coupled to a top end of the bottom platen side wall32may define a bottom platen engagement surface40. The bottom platen engagement surface40may extend around a perimeter of the bottom platen base receptacle36, thereby to provide a first surface for securing the outer edge of the tissue sample26. The bottom platen engagement surface40is spaced from the bottom platen base inner surface34to limit the amount of structure in direct contact with the tissue sample26. To provide a stable surface on which the biological material may be placed, the bottom platen engagement surface40may be substantially planar.

Turning toFIG. 4, the top platen24is illustrated as having a generally inverted, tray-like appearance formed by a top platen cover42and a top platen side wall44. The top platen cover42defines a top platen cover inner surface46, while the top platen side wall44extends downwardly from a perimeter of the top platen cover42. Together, the top platen side wall44and top platen cover42define a top platen cover receptacle48. As best shown inFIG. 3, a bottom portion of the top platen side wall44defines a top platen engagement surface50. The top platen engagement surface50extends around a perimeter of the top platen cover receptacle48, thereby to provide a second surface for securing the outer edge of the tissue sample26. The top platen engagement surface50is spaced from the top platen cover inner surface46to again limit direct contact with the tissue sample26.

The top platen engagement surface50is shaped to cooperatively interact with the bottom platen engagement surface40. As used herein, the term “cooperatively interact” means that the recited structures are configured so that they can be positioned in a tissue engaging position, thereby to secure the tissue sample26between the two engagement surfaces. Accordingly, in some embodiments, the top platen engagement surface50may have a shape that is a mirror image of that of the bottom platen engagement surface40, so that the engagement surfaces40,50can be directly aligned. In these embodiments, the engagement surfaces40,50may be configured to directly abut one another, or they may be configured to be spaced by a distance that is sufficiently small to firmly secure the tissue sample26therebetween. Alternatively, the engagement surfaces40,50may be configured to have a closely telescoping fit, thereby to pinch an outer edge of the tissue sample26therebetween. The engagement surfaces40,50may be configured to sever, shear, or otherwise separate an outer periphery of the biological material to obtain the final shape of the tissue sample26. Alternatively, the engagement surfaces may simply secure the tissue sample and a separate cutting step may be performed to remove the excess periphery of the biological material.

The bottom and top platens22,24may be provided as separate components that can be assembled to secure the tissue sample26. To assist with assembly, the bottom and top platens22,24may include alignment components so that the top platen engagement surface50is positioned properly with respect to the bottom platen engagement surface40. As shown inFIG. 2, the bottom platen22may include a first alignment structure, such as two pairs of brackets52extending outwardly from opposite ends of the bottom platen side wall32, while the top platen24may include a second alignment structure, such as two tabs54extending downwardly from opposite ends of the top platen side wall44that are sized for insertion between the two pairs of brackets52. The bracket pairs52and tabs54are located such that when the tabs54are inserted into the respective bracket pairs52, the top platen engagement surface50will be properly positioned relative to the bottom platen engagement surface40, thereby permitting cooperative interaction therebetween.

The tissue securing device20may further include a coupler assembly configured to releasably couple the bottom and top platens22,24. As best shown with reference toFIGS. 1 and 3, the coupler assembly may include a latch56having a base rotatably coupled to the top platen24and a hook end58. Additionally, the bottom platen22may be formed with a coupler recess60configured to releasably engage the hook end58of the latch56. Accordingly, when the top platen24is assembled on top of the bottom platen22, the latch56may be rotated until the hook end58engages the coupler recess60, thereby to firmly secure the top platen24to the bottom platen22. As the coupler assembly is locked in place, the top platen24may be further pulled toward the bottom platen22, driving the top platen engagement surface50toward the bottom platen engagement surface40and further securing the periphery of the tissue sample26.

The tissue securing device20may be configured to promote exposure of the tissue sample26to controlled exterior environments, such as transport fluid or cryogenic media. As best shown inFIGS. 2 and 3, the bottom platen base30defines a plurality of bottom platen base apertures62that fluidly communicating between an exterior of the bottom platen22and the bottom platen base receptacle36. Additionally, as best shown inFIGS. 3 and 4, the top platen cover42defines a plurality of top platen cover apertures64fluidly communicating between an exterior of the top platen24and the top platen cover receptacle48. The bottom platen base apertures62and top platen cover apertures64promote fluid communication from an exterior of the securing device20and the tissue sample26, which can be beneficial when the device20is submerged in transport fluid or exposed to cryogenic media.

The bottom and top platens22,24of the securing device20may be used on their own to obtain a relatively large tissue sample26. In operation, the top platen24is initially separated from the bottom platen22. Biological material may be placed over the bottom platen22so that an outer periphery of the biological material drapes over the sides of the bottom platen22(FIG. 5). Using the alignment components, the top platen24may be assembled onto the bottom platen22until the top platen engagement surface50is properly positioned relative to the bottom platen engagement surface40(FIG. 6). Next, the coupler assembly may be engaged to lock the top platen24onto the bottom platen22(FIG. 7). During this process, the top platen engagement surface50is moved into a tissue engaging position relative to the bottom platen engagement surface40, so that a perimeter portion of what is to be the tissue sample26is firmly secured therebetween. The excess periphery of biological material is removed either automatically or in a separate cutting process, thereby to obtain the final shape of the tissue sample26(FIG. 1).

While the bottom and top platens22,24have separate utility, the securing device20may further include an array of diskette assemblies70for separating the tissue sample26into multiple, smaller sub-samples. Each diskette assembly70may include a diskette lower housing72and a diskette upper housing74, as best shown inFIGS. 8 and 9.

The diskette lower housing72may include a lower housing base76defining a lower housing base inner surface78. A lower housing side wall80extends upwardly from the lower housing base76so that the lower housing base76and the lower housing side wall80define a lower housing receptacle82. A lower housing shoulder84projects inwardly from the lower housing side wall80to define a lower housing engagement surface86. The lower housing engagement surface86extends around a perimeter of the lower housing receptacle82and is spaced a distance above the lower housing base inner surface78.

The diskette upper housing74may include an upper housing cover88defining an upper housing cover inner surface90. An upper housing side wall92extends downwardly from the upper housing cover88so that the upper housing side wall92and upper housing cover88define an upper housing receptacle94. The upper housing side wall92has a lower edge96defining an upper housing engagement surface98. The upper housing engagement surface98extends around a perimeter of the upper housing receptacle94and is spaced a distance below the upper housing cover inner surface90. The upper housing engagement surface98may be shaped to cooperatively interact with the lower housing engagement surface86, similar to the engagement surfaces40,50described above.

The bottom platen22may be configured to secure the diskette lower housings72in defined locations to facilitate registration of each diskette lower housing72with an associated diskette upper housing74. As best shown inFIG. 1, the bottom platen base inner surface34defines an array of bottom platen base recesses100, each of which is sized to receive a diskette lower housing72. The bottom platen base recesses100may be formed by a series of intersecting platforms102that are elevated from the bottom platen base inner surface34, but not as high as the bottom platen engagement surface40. A base retainer104may extend upwardly from the bottom platen base inner surface34into each bottom platen base recess100and may be configured to releasably attach to a retainer aperture106formed in the diskette lower housing72, as best shown inFIG. 10.

Similarly, the top platen24may be configured to secure the diskette upper housings74in defined locations to facilitate registration with an associated diskette lower housing72. As best shown inFIG. 4, the top platen cover defines an array of cover apertures64. Each upper housing cover88is sized for insertion through the cover apertures64. An upper housing shoulder108(FIG. 8) extends outwardly from the upper housing cover88and defines a shoulder surface110configured to engage the top platen cover inner surface46, thereby to prevent further insertion of the diskette upper housing74through the cover aperture64. A flexible cover tab109may project outwardly from the upper housing cover88that may deflect to pass through the cover aperture64. Once the cover tab109is clear of the cover aperture64, it may resume its original shape, thereby by to secure the diskette upper housing74in a normal position. Subsequently, when sufficient downward force is applied to the upper housing cover88, the cover tab109may again deflect to permit the diskette upper housing74to move downwardly to an actuated position, as discussed in greater detail below.

Each cover aperture64is positioned to register with an associated bottom platen base recess100, so that each associated cover aperture64and bottom platen base recess100define a diskette receptacle configured to ultimately receive and enclose a diskette assembly70. For example, bottom platen base recesses100and cover apertures64may be located relative to the alignment components of the bottom and top platens22,24, respectively, such that when the alignment components are engaged to assemble the bottom and top platens22,24, the lower and upper housing engagement surfaces86,98are simultaneously aligned.

Each diskette assembly70may be configured so that the diskette lower housing72automatically engages the diskette upper housing74during operation, such as when the diskette upper housing74is dislodged from the top platen24and moved to the actuated position. For example, as best shown inFIG. 12, the diskette lower housing72may include a lower housing retainer in the form of a lower housing flange111flange extending inwardly from the lower housing side wall80and defining a lower housing flange surface112. The diskette upper housing74may include an upper housing retainer in the form of a flexible tab114protruding outwardly from the upper housing side wall92and defining an upper tab surface116. As the diskette upper housing74is driven toward the diskette lower housing72, the tab114may deflect until it passes the lower housing flange111, at which time the tab114may resume its original shape. In this position, the upper tab surface116will engage the lower housing flange surface112to resist separation of the diskette upper housing74from the diskette lower housing72.

Movement of the diskette upper housing74from the normal position to the actuated position may facilitate separating one or more sub-samples120from the tissue sample26. In the normal position, where the diskette upper housing74engages the top platen22, the upper housing engagement surface98is positioned above the top platen engagement surface50and is aligned with but spaced from the lower housing engagement surface86, as shown inFIG. 12. In the actuated position, the diskette upper housing74is disengaged from the top platen22and moved downwardly so that the upper housing engagement surface98is positioned below the top platen engagement surface50, as best shown inFIG. 3. When in the actuated position, the upper housing engagement surface98may be in a tissue engaging position such that a periphery of the sub-sample120is pinched between the lower and upper housing engagement surfaces86,98(FIG. 3). The engagement surfaces86,98may abut or may be spaced by a distance sufficiently small so that the sub-sample120is secured therebetween. In some embodiments, the lower and upper housing side walls80,92may be configured to shear, cut, or otherwise sever through the tissue sample26.

Each diskette assembly70may be configured to promote fluid communication from the exterior environment to an interior space surrounding the tissue sub-sample120. As best shown inFIGS. 8-12, each upper housing cover88defines a plurality of upper housing cover passages130fluidly communicating between an exterior of the upper housing cover88and the upper housing receptacle94. Additionally, each lower housing base76defines a plurality of lower housing base apertures132fluidly communicating between an exterior of the lower housing base76and the lower housing receptacle82. A bottom surface134of the diskette lower housing72may further be formed with channels136extending between each of the lower housing base apertures132, thereby to further promote fluid flow into the lower housing receptacle82. Additionally, the upper housing engagement surface98may be configured to promote additional fluid flow to the sub-sample120. More specifically, the upper housing engagement surface98may be formed with a series of upper housing engagement surface passages138(FIG. 12) that fluidly communicate between an exterior of the diskette upper housing74and the upper housing receptacle94.

In some embodiments, the device20may be configured to facilitate a separate cutting step to separate the sub-sample120. In these embodiments, the sub-sample120is not separated simultaneously as the diskette upper housing74is moved to the actuated position. Instead, movement of the diskette upper housing74only secures a periphery of the sub-sample120between the lower and upper housing engagement surfaces86,98. A separate cutting step may be facilitated by sufficiently spacing the bottom platen base recesses100so that a cutting gap142is provided between lower housing side wall peripheries140of adjacent diskette lower housings74(FIG. 3). The platforms102may extend between the adjacent bottom platen base recesses100, and therefore an upper platform surface144may be aligned with the cutting gap142to provide stop for the cutting blade, as shown inFIG. 13. An individual diskette assembly70holding a sub-sample120may then be separated from the bottom platen22, as shown inFIG. 14.

A diskette assembly70having engaged diskette lower and upper housings72,74is illustrated inFIGS. 15-17. From these drawings it will be appreciated that a periphery of the sub-sample120is secured within the housings72,74with minimal direct contact. Additionally, it is seen how the upper housing cover passages130, lower housing base apertures132, channels136, and upper housing engagement surface passages138promote fluid communication between an exterior of the diskette assembly70and the sub-sample120.

FIG. 18illustrates two devices20stacked in a nested configuration. The stacked devices20may be inserted into a transport vessel150as shown inFIG. 19. The transport vessel150may be filled with a preservative solution, which may intimately contact the tissue samples26through the various apertures and passages in the platens22,24and diskette housings72,74noted above. When the devices20arrive at the intended destination, such as a cryogenic storage facility, the entire device20holding the full tissue sample26may be stored, or individual sub-samples120housed in diskette assemblies70may be separated and stored as needed. If the entire device20is cryogenically stored and only a sub-sample120is later needed, the device20may be thawed and the sub-sample120separated without direct handling of the tissue sample26. Similarly, the device20permits an explant culture procedure for stem cell derivation to be performed, and is capable of being immersed in a cryogenic preservation medium and returned to cryogenic storage more than one time without directly handling the tissue sample.

FIG. 20illustrates a diskette assembly70disposed in an individual cryogenic vial160. The cryogenic vial160may be filled with a preservative solution and prepared for cryogenic storage, thereby allowing individual sub-samples120to be separated and stored with minimal handling of the tissue.

INDUSTRIAL APPLICABILITY

The device as described allows a placental tissue sample to be secured, transported in an appropriate protection medium, transferred to an appropriate cryogenic preservation medium and frozen intact. The device further allows, at a future time, a thawing procedure followed by an explant stem cell derivation culture procedure, which upon conclusion allows the return to cryogenic preservation for a second or more times.

The device may be used to capture a tissue sample at a child birthing event; permit immersion in a medium for transportation preservation to the next laboratory site; permit transfer from a transportation medium to a cryo-preservation medium; permit cryogenic storage; permit removal and thawing from cryogenic storage; and permit subsequent explant culturing to derive Mesenchymal-like stem cells, all with minimal or no direct handling of the sample. Furthermore, the device allows the sample to sustain repetitive cycles of cryogenic freezing, thawing, and explant stem cell derivation.

Additionally, portions of the device may be constructed of a biomimetic material that can serve as a tissue scaffold that can be directly utilized after a placental tissue derived Mesenchymal-type stem cell derivation procedure.

In view of the foregoing, the subject matter disclosed herein may facilitate one or more of the following advantages:

(1) Providing a staging platform or work surface upon which to present and stabilize a large and unwieldy tissue sample for further manipulation and preparation. This is accomplished by a rigid bottom platen assembly. Optional detachable legs are offered as a means of additional elevation of the working surface as needed;

(2) After staging, the sample may be secured in a manner that retains flatness and prevents folding, curling, shrinkage or other movement, as the sample will normally tend to migrate in many directions. This is accomplished by mating a rigid top platen assembly over the bottom platen assembly, and having the ability to clamp the two platen halves together, thus capturing the large tissue sample between the halves.

(3) Because a singular large sample may be most conducive to transportation for further professional preservation, the platen assemblies may be configured to permit nested stacking in a compact manner that facilitates insertion into a transport container. Also, because exposure to air should be limited, the large tissue sample may be quickly placed within a preserving liquid solution, and the components comprising the platen assembly may facilitate maximum intimate liquid contact with top and bottom tissue surfaces when the platens are immersed in the solution. Discrete passages may be formed in the surfaces contacting and retaining the tissue sample to promote fluid contact with the tissue sample. In some embodiments, the platen materials in contact with the tissue sample may be formed of a hydrophilic material to maximize surface wetting.

(4) The ability to separate the large tissue sample into multiple sub-samples may provide additional advantages. When tissue samples are cryogenically preserved, a common practice is to thaw the entire tissue sample, extract the desired sub-sample, and then re-freeze the remainder of the tissue sample. Each thaw and freeze cycle is believed to diminish the viability of the sample. Accordingly, it may be advantageous to provide the ability to freeze and thaw smaller pieces of the large sample on a more selective basis, as permitted by the use of the diskette assemblies taught herein.

(5) Similar to transport conditions, when the large tissue sample is transformed into smaller multiple sub-samples for cryogenic storage, the sub-samples are typically immersed in a cryogenic preservation medium. The methods and apparatus disclosed herein promote intimate contact between the surfaces of the tissue sample and the preservation medium.