Methods of using container system to reduce abrasion or other types of damage to tissue during tissue processing, methods of reducing abrasion or other types of damage to tissue during tissue processing, and tissue specimens processed by methods thereof

Abstract

To facilitate multiple steps of tissue processing.SOLUTION: A tissue processing container is specifically designed to achieve automated tissue processing, exposure of tissues to processing media, and efficiency of multi-step tissue processing protocols. The described tissue processing containers include a cup-like cavity with a central hole covered by a mesh screen, a trough for separate processing steps, and are stackable in a substantially air-tight and substantially water-tight manner. Tissues for processing using the described tissue processing containers include neural tissue. A transport housing for transporting stacks of the tissue processing containers is also described herein, which may directly incubate the containers, or the transport housing may be placed inside a separate incubator for incubating the containers.SELECTED DRAWING: Figure 1A

Description

[Technical field]

[0001] The present invention relates to a tissue processing container for automated processing of tissue, a method of use thereof, and a system including the tissue processing container. [Background technology]

[0002] Tissues such as skin tissue, vascular tissue, cardiac tissue, nervous tissue, umbilical cord tissue, and the like, are useful in various types of surgery and wound healing in humans and animals. Tissues used in surgery and / or transplantation procedures may be harvested from humans or animals. Some tissues, such as skin tissue, may also be artificially produced for testing and use. Examples of commercially available processed tissues include Avance® nerve grafts and Avive® soft tissue membranes manufactured by Axogen, Inc. (Alachua, Florida, USA).

[0003] Processing, including but not limited to decellularization or reduction of bioburden of non-autologous tissues for surgical implantation, can be time-consuming and laborious, particularly in terms of the many different solutions, enzymes, cleaners / rinses, or other liquid media to which the tissue may be exposed. Tissue processing can result in unacceptable abrasions or even other types of damage to the tissue, compromising the surgical outcome of the implantation and adversely affecting the integrity of the tissue and prohibiting its ultimate surgical use.

[0004] Thus, there is a need for more efficient and tissue-friendly methods of tissue processing (eg, methods that better protect the integrity of the tissue and its structures). Summary of the Invention

[0005] More automated means, including tissue processing containers, useful for facilitating multiple steps of tissue processing, and components therefor, are described herein. The tissue processing containers may be capable of stacking, including in a substantially air-tight and substantially liquid-tight manner, each capable of processing one or more tissue specimens per container. Systems incorporating the tissue processing containers are also disclosed. Transport housings for the tissue processing containers are further disclosed. The tissue processing containers, systems, and general methods described herein may be used to expose tissues to various liquid media, which may be beneficial, for example, for decellularization or reduction of bioburden, among other tissue processing efforts.

[0006] For example, a tissue processing container is provided that is adapted for automation and multiple steps of tissue processing. The container may possess a circular interior shape, but an exterior shape that may be of any design or dimensions, for example, adaptable for robotic manipulation and processing. The container may have the appearance of a full cup, with a second inner diameter and a hole in the center of the container through which air and liquid can pass. The hole may be substantially circular, oval or elliptical in shape, or may be polygonal in shape. The outer wall, i.e., the first wall, of the tissue processing container may be of any height, and in some embodiments may be perpendicular and straight from the circular bottom of the container, and in other embodiments may be slanted or flared outward from the center of the container, and / or may not be perpendicular from the circular bottom of the container.

[0007] In some embodiments, the second wall may surround a hole in the circular bottom of the container. The height of the second wall may be less than the height of the first wall. When placed on its circular bottom, the tissue processing container may be open-topped like a cup. The tissue processing container may have one or more troughs rigidly attached and / or integrally formed in the circular bottom of the tissue processing container and surrounding the hole. The tissue processing container may hold a first volume of liquid and the one or more troughs may hold a second volume of liquid, which may be more than the second volume of liquid.

[0008] In some embodiments, the tissue processing container may further possess two mesh screens and two gaskets. The first mesh screen may be rigidly attached to a hole on the inner surface of the tissue processing container. The first gasket may surround a second wall surrounding the hole on the inner surface of the tissue processing container. The second gasket may be attachable to the outer surface of the circular bottom of the tissue processing container and may have a diameter approximately equal to the second diameter. The second mesh screen may be attachable to the outer surface of the circular bottom. The two mesh screens may be made of plastic polymers, pure or synthetic metals, resins, and / or elastic polymers such as rubber, or combinations thereof. In specific embodiments, the first and / or second mesh screens may be comprised of polyamides such as nylon. In further embodiments, the first and / or second mesh screens may be comprised of nylon and polypropylene. The gauge or thread count of the mesh may be sufficient such that any contact between the mesh and the tissue does not cause unacceptable abrasion or damage to the tissue during tissue processing. A sufficient gauge or thread count may be about 300X300 threads per inch, and about 500X500 threads per inch. For example, a suitable gauge or thread count for the mesh may be about 325X325 threads per inch, or a suitable mesh may include openings of less than about 0.0017 inches.

[0009] In certain embodiments, the second mesh screen, when attached or coupled to the exterior side of the circular bottom of the tissue processing vessel, may not physically touch or be associated with the exterior side of the circular bottom of the tissue processing vessel, i.e., the second mesh screen may be spaced apart from the exterior side of the circular bottom of the tissue processing vessel.

[0010] In one embodiment, when the tissue processing container is placed on its circular bottom on a flat surface with its top open upward, the highest features, in order of height from top to bottom, may be the first wall, followed by the first mesh screen, the first gasket, the second wall, one or more troughs, the circular bottom, the second mesh screen, and the second gasket, for example, as depicted in FIG. 1B.

[0011] In another embodiment contemplated herein, the tissue processing container may be stackable with one or more additional tissue processing containers, and when the tissue processing containers are stacked, the central holes of the containers in the stack may align. The central holes of the tissue processing containers in the stack may be of the same shape as one another, e.g., all circular, all octagonal, etc. In such an embodiment, the second gasket may form a substantially air-tight and substantially water-tight seal between the two tissue processing containers such that neither air nor liquid can substantially escape from between the two tissue processing containers when filled.

[0012] In some embodiments, the one or more troughs may be adjacent to and touching the first wall in a specific embodiment. In another embodiment, the one or more troughs may be adjacent to and touching the second wall. In further embodiments, the one or more troughs may not touch the first wall. In some embodiments, the one or more troughs may be semicircular in shape. In one embodiment, the one or more troughs may each be large enough to accommodate at least one tissue specimen.

[0013] In one embodiment, the tissue processing container may be made from a polymer or plastic polymer, such as, but not limited to, one or more of polycarbonate, polyetherimide, polyphenylsulfone, polystyrene, polysulfone, and polymethylpentene, and the like. In a specific embodiment, the tissue processing container may be made from polymethylpentene (also known as NALGENE®, manufactured by Thermo Fisher Scientific, Waltham, Massachusetts, USA). The tissue processing container may alternatively be constructed from glass or silicate. In some embodiments, the tissue specimen may not substantially adhere to the surface of the tissue processing container.

[0014] In certain embodiments, the inner surface of the outer side of the circular bottom of the tissue processing container may be sloped from the outer diameter to the inner diameter, i.e., from the second diameter to the first diameter, towards the central hole so that when the tissue processing container is oriented so that the bottom is over the trough, i.e., so that the cup of the tissue processing container is oriented upside down, all liquid contacting the outer side drains through the central hole.

[0015] In a specific embodiment, the first volume may be between about 100 mL and 500 mL. In another such embodiment, the second volume may be between about 1 mL and 20 mL. In some embodiments, the first wall may include a notch etched or affixed thereto to indicate a fill level when liquid is added to the tissue processing container.

[0016] As noted above, in certain embodiments, the first wall may have a thickness of about 1-5 mm. In other embodiments, the tissue processing container may include an external second container such that the external dimensions of the tissue processing container are different from the internal dimensions of the tissue processing container. In such embodiments, the second container may be of a larger shape, which may be, for example, circular, oval, elliptical, or the like, or polygonal. The shape of the second container may facilitate handling or gripping of the tissue processing container, including handling or gripping by robotic instruments and automated machines. In one such embodiment, the second container may have a height approximately equal to the height of the first wall. The second container may be constructed of one or more of polycarbonate, polyetherimide, polyphenylsulfone, polystyrene, polysulfone, and polymethylpentene, and the like, or glass or silicate.

[0017] In certain embodiments, the height of the first wall may be from about 1 cm to about 10 cm. In alternative embodiments, the tissue processing container may further include a lid having a diameter approximately equal to the second diameter.

[0018] In further embodiments of the described tissue processing containers, two or more tissue processing containers may be stacked on top of each other, with at least one tissue processing container being a bottom tissue processing container and positioned below all of the other tissue processing containers in the stack, and at least one tissue processing container being a top tissue processing container positioned above all of the other tissue processing containers in the stack. Optionally, the tissue processing containers may each include a sloped outer surface such that the outer edge of the bottom of the tissue processing container extends further than the outer edge of the top of that same tissue processing container, and when stacked, the outer edge of the bottom of the tissue processing container extends beyond the outer edge of the top of the tissue processing container next to it. In some embodiments, there may additionally be a base container positioned below the bottom tissue processing container. The base container may possess substantially the same internal dimensions as the bottom tissue processing container, such that the bottom tissue processing container fits into the base container in a manner similar to how one tissue processing container fits into another tissue processing container when stacked. The base container may further include a first nozzle attached to the first wall of the base container and extending into an interior side of the base container. Optionally, the first nozzle extending into the interior side of the base container may further couple to a central bore of an adjacent tissue processing container.

[0019] In such stacked tissue processing container embodiments, in some embodiments, there may also be a cap container possessing substantially the same internal dimensions as the top tissue processing container, such that the cap container fits with the top tissue processing container in a manner similar to how one tissue processing container fits into another tissue processing container when stacked. The cap container may further include a second nozzle attached to the first wall of the cap container and extending into the interior side of the cap container. Optionally, the second nozzle extending into the interior side of the cap container may further directly connect to the central hole of an adjacent tissue processing container. In the aforementioned embodiment, each of the central holes of all the tissue processing containers may be aligned, and further a substantially air-tight and substantially water-tight seal may be formed between each tissue processing container. In some embodiments, a substantially air-tight and substantially water-tight seal may also be formed between the bottom tissue processing container and the base container, and also between the top tissue processing container and the cap container when the base container and the cap container are included in the stack.

[0020] In certain embodiments, the cap container and the base container may not themselves be configured or used for processing tissue, for example, the cap container and / or the base container may be used for fluid management.

[0021] Some embodiments of the stacked tissue processing container may also include a cap container and a base container, and the first and second nozzles may each be attached to a hose that may direct air and / or liquid therethrough.

[0022] In other embodiments, when in use, each tissue processing container of the stacked tissue processing containers may contain one or more tissue specimens, which may be in contact with the second circular mesh screen of the tissue processing container, and the one or more tissue specimens within the container are present within the tissue processing container.

[0023] In some embodiments, the base container and the cap container may be identical and each may possess two nozzles. In alternative embodiments, such cap and base containers may further possess a slanted disk that bisects the first wall of the cap and base container, so that the first nozzle may be located on or above the slanted disk and the second nozzle may be located below the slanted disk in such a way that one may be placed on top of the stack of tissue processing containers and the other may be placed below the stack of tissue processing containers depending on how the cap and base containers are oriented, and liquid and / or gas may flow through the respective nozzles as desired. In such embodiments, the first and second nozzles may be on opposite sides of each other in the first wall and may not contact the central hole of the bottom or top tissue processing container.

[0024] Also described herein are tissue processing systems including one or more stacks of tissue processing containers, each stack optionally including both a cap container and a base container as described herein, and further including a transport housing that encases the one or more stacks of tissue processing containers. Such transport housings may be further sealed and / or temperature controlled to provide desired conditions for a particular tissue processing step (such as, but not limited to, incubation at a precise temperature without the need for an incubator separate from the transport housing).

[0025] The tissue processing system may, in some embodiments, also include a programmable controller unit adapted to control various time-dependent external inputs and hardware, such as mechanical / electrical pumps that drive the flow of fluids and air into and out of the stacked tissue processing containers, as well as temperature controllers and other control means for regulating the vibration speed of the tissue processing containers, for example via one or more shaker plates on which the stacked tissue processing containers may be mounted.

[0026] The tissue processing system may further include additional external hardware elements controlled by the controller, such as an incubator including a compartment large enough to accommodate the transport housing, so long as the housing itself is not capable of controlling the temperature of the tissue processing containers stacked therein. At least two hoses possessing diameters approximately equal to those of the first and second nozzles are also contemplated as part of such a system, for transferring necessary fluids and gases to and from the stack of tissue processing containers.

[0027] As described, in some embodiments, the system may include one or more hose-mounted electric pumps that are electronically controlled by the controller unit and adapted to rigidly couple to at least one of the at least two hoses.

[0028] The hoses in such systems may in turn be attached to one or more media reservoirs containing tissue processing media and other solutions beneficial to the processing of tissue. Also contemplated as part of such systems are one or more waste units adapted to be attached to one of the at least two hoses for the purpose of facilitating evacuation of liquids and gases from the tissue processing container.

[0029] Another contemplated external hardware element electrically controlled by the controller unit is a shaker that includes a surface adapted to receive the housing or fit inside the housing such that the tissue processing container vibrates at a desired frequency to maximize contact of the tissue specimen with the surrounding medium.

[0030] Methods of processing tissue are also provided herein. According to one tissue processing method of the present disclosure, a tissue processing container is oriented such that the interior side of the bottom faces upward. A tissue specimen may be placed in a trough of the tissue processing container. A first tissue processing medium may include one or more of an enzyme, detergent, salt solution, or other liquid medium, and may be added to the trough. The processing medium may be added to the trough before, after, or at the same time that the tissue specimen is placed in the trough. These steps may be repeated with additional tissue processing containers, forming a stack of tissue processing containers containing tissue specimens and the first processing medium. An individual tissue processing container, or a stack of such tissue processing containers, containing tissue and the first tissue processing medium may be incubated at a first temperature for a first period of time. The stack may or may not also include a cap container and a base container. The medium of the tissue processing container may then be removed by turning the stack of tissue processing containers upside down, thereby draining the first tissue processing medium from the tissue processing container. The tissue specimens in the stack of tissue processing vessels may then be rinsed / washed / soaked / or otherwise exposed to and possibly cultured in a second, third, fourth, ... or nth additional processing medium until the tissue processing method is completed. In doing so, the stack of tissue processing vessels may be placed on a base vessel including a nozzle, and a cap vessel including a nozzle may be placed on top of the stack of tissue processing vessels. A second tissue processing medium (which may include a fresh aliquot of the first tissue processing medium, or alternatively a different tissue processing medium) may be filled through the nozzle of the bottom-most vessel. Optionally, the second tissue processing medium (or any additional tissue processing medium used to process the tissue in further processing steps) may be added to the tissue processing vessel through the nozzle of the top-most vessel.For example, when filling through the nozzle of the bottommost vessel (the bottommost vessel being the cap vessel located on the bottom of the stack when the stack is inverted to drain the first tissue processing medium) until the fluid level of the tissue processing medium reaches the second wall of the central hole of the bottommost tissue processing vessel in the stack, the second tissue processing medium may stop filling the bottommost tissue processing vessel in the stack when its fluid level contacts the second wall of the central hole of the bottommost tissue processing vessel in the stack, and any additional second tissue processing medium injected or otherwise filled into the bottommost tissue processing vessel in the stack may exit through the central hole and enter the tissue processing vessel stacked on top of the bottommost tissue processing vessel in the stack, and so forth, whereby the second tissue processing medium fills all of the tissue processing vessels in the stack. In certain embodiments, the first tissue processing medium comprises at least one active enzyme, and in other embodiments, the first tissue processing medium comprises multiple active enzymes.

[0031] In some embodiments, such methods may also include placing the tissue processing container containing the second tissue processing medium in an incubator, and optionally incubating the tissue processing container at a second temperature (which may or may not be the same as the first temperature) for a second period of time (which may or may not be the same period of time as the first period). In certain embodiments, the first and second periods of time may not be identical. In other embodiments, the tissue processing container (with or without an incubator) may be placed on a shaker. When used, the shaker may be operated for a period of time sufficient to generate standing waves within the tissue processing container. In additional embodiments, the tissue may contact the second mesh screen when the stack of tissue processing containers is turned upside down.

[0032] The above method may, in additional embodiments, include additional steps such as draining the second tissue processing medium from all of the tissue processing containers, which may pass through a central hole and exit through a nozzle in the cap container (which will be placed at the bottom of the stack of tissue processing containers if the cap container was included in the stack before the stack was inverted). It should be recognized that the terms "cap container" and "base container" may be used interchangeably and both refer to a single end container depending on when that container was added to the stack of tissue processing containers. That is, the container placed on top of the stack of tissue processing containers before the tissue processing container is inverted would be referred to as the cap container and would be on the bottom of the stack after the tissue processing container is inverted. The container placed on top of the stack of tissue processing containers after the tissue processing container is inverted would be referred to as the cap container and would be on top of the inverted stack.

[0033] Aspects of the present disclosure may be directed to a container system. The container system may include a cup having an outer wall and an inner wall disposed radially inward of the outer wall, and the height of the inner wall may be less than the height of the outer wall. The system may also include a floor (also referred to as a "bottom" in this disclosure) of the cup, the floor having a hole substantially in the center of the floor, the hole being disposed radially inward of the inner wall such that the inner wall surrounds the hole and the outer wall surrounds the inner wall. The system may further include at least two troughs disposed on an inner side of the floor of the cup, the at least two troughs surrounding the hole and the at least two troughs being defined by two or more trough walls protruding from the inner side of the floor.

[0034] Various aspects may further include one or more of: a mesh screen covering the hole; a gasket configured to tightly seal the mesh screen against the inner wall; a mesh screen disposed on an exterior side of the floor opposite the interior side; a gasket configured to seal the mesh screen against the exterior side of the floor or the exterior side of the outer wall; the exterior wall may extend radially outward such that the exterior wall extends away from the floor of the cup; the floor may be sloped such that the floor extends from the exterior wall to the inner wall; an end of the cup opposite the floor may be uncovered; or the substantial center of the floor may be the center of the floor.

[0035] Another aspect of the disclosure may be directed to a container system including a cup having a perimeter defined by an outer wall, the cup having a closed end and an open end, and an inner wall disposed radially inward of the outer wall, the inner wall having a height less than the height of the outer wall. The system may further include a floor defining the closed end of the cup, and a hole disposed substantially in the center of the floor, the hole disposed radially inward of the inner wall such that the inner wall surrounds the hole and the outer wall surrounds the inner wall. The system may also include at least two troughs disposed on an interior surface of the floor of the cup, the at least two troughs being defined by two or more trough walls projecting from the floor toward the open end of the cup, each of the two or more trough walls having a height less than the height of the inner wall, a first mesh screen extending across the hole, and a second mesh screen extending along an exterior surface of the closed end of the cup.

[0036] Various embodiments may further include one or more of: a first gasket configured to tightly seal the first mesh screen across the hole, and a second gasket configured to seal the second mesh screen to the closed end of the cup; an end of the outer wall closest to the floor of the cup may define a first diameter, an end of the inner wall closest to the floor of the cup may define a second diameter smaller than the first diameter, and the hole may define a third diameter smaller than the second diameter; the floor of the cup may slope radially outward from the second diameter to the first diameter such that the floor adjacent the first diameter is located farther from the open end of the cup than the floor adjacent the second diameter; a lid configured to removably cover the open end of the cup; or a substantial center of the floor may be a center of the floor.

[0037] A further aspect of the present disclosure may be directed to a container system including a first cup and a second cup, each of the first cup and the second cup including an open end and a closed end; an outer wall extending from the open end to the closed end and defining a perimeter; an inner wall disposed radially inward of the outer wall, the inner wall having a height less than the height of the outer wall; a floor having a hole substantially in a center of the floor, the hole being disposed radially inward of the inner wall such that the inner wall surrounds the hole and the outer wall surrounds the inner wall; and at least two troughs disposed on an inner surface of the floor of the cup, the at least two troughs being defined by two or more trough walls protruding from the inner surface of the floor. Further, the closed end of the first cup may be dimensioned to be received within the open end of the second cup such that the first cup is stackable with the second cup, and the hole in the first cup aligns with the hole in the second cup when the first cup is stacked with the second cup.

[0038] Various aspects include that each of the first cup and the second cup may further include a mesh screen extending across the hole, each of the first cup and the second cup may further include a gasket that securely fastens the mesh screen across the hole, each of the first cup and the second cup may further include a second mesh screen extending along an exterior surface of the closed end of the cup, each of the first cup and the second cup may further include a second gasket that secures the second mesh screen along an exterior surface of the closed end of the cup, an outer wall of the first cup and The outer walls of the two cups may each define a first diameter adjacent the closed end that is smaller than a second diameter adjacent the open end, and the system may further include a base container having a first nozzle extending into an interior region of the base container, the first nozzle configured to be fluidly connected to the hole in the first cup, and a cap container having a second nozzle extending into an interior region of the cap container, the second nozzle configured to be fluidly connected to the hole in the second cup, or the substantial center of the floor may be the center of the floor.

[0039] This Summary of the Invention is provided to guide the selection of the subject matter that is further described in the Detailed Description below. This Summary is not intended to identify key or essential features of the claimed subject matter, nor is it intended to limit the scope of the claimed subject matter, which is more fully described herein below.

[0040] Reference is made herein to specific embodiments as illustrated in the figures. The figures are not drawn to scale, and any reference to dimensions in the figures or in the following description is to specific embodiments. It will be apparent to one of ordinary skill in the art that variations in these dimensions are possible while still maintaining full functionality for the intended purpose. Such variations are specifically contemplated and incorporated into the present disclosure, regardless of the specific embodiments depicted in the figures below. [Brief description of the drawings]

[0041] [Figure 1A] FIG. 2 is an exploded view of the tissue processing container. [Figure 1B] FIG. 2 is a side view of the tissue processing container. [Figure 2A] FIG. 13 is a side view of a stack of tissue processing containers including a base container with a nozzle and a cap container. [Figure 2B] FIG. 13 is a perspective view of an alternative embodiment cap / base container. [Figure 3A] FIG. 1 illustrates a system including stacking of tissue processing containers. [Figure 3B] FIG. 1 illustrates a system including stacking of tissue processing containers. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0042] definition The term "gasket" means a ring-shaped object, made, for example, from a plastic polymer, a rubber polymer, a resin, or a mixture thereof, which tightly engages opposite surfaces and acts to create a substantially air-tight and / or substantially water-tight seal between the surfaces when pressure is applied thereon.

[0043] The term "trough" refers to an open vessel of any dimension or size that is firmly attached to, incorporated into, integrally formed with, or otherwise formed as part of the surface of a tissue processing vessel as described herein and shown, for example, in Figures 1 and 2. The trough is open to allow for the addition or removal of liquids and other materials that enter or leave the trough.

[0044] The term "tissue" refers to nervous tissue or other types of tissue such as, but not limited to, heart, muscle, tendon, ligament, blood vessel, skin, SIS (small intestine submucosa), umbilical cord, dura mater, fascia, serosa, periosteum, and other types of tissue. The tissue may be obtained from human or animal sources and can be, but is not limited to, rodent, equine, canine, leporid, porcine, primate, ovine, or other ruminant, or other derived. The tissue may be naturally obtained from a human or animal source, or the tissue may be grown in a laboratory.

[0045] The term "control unit" or "controller" refers to power hardware devices including, for example, a controller or microcontroller, a processor or microprocessor with associated memory and peripherals, a field programmable gate array (FPGA), a programmable logic array (PLA), or a programmable logic controller (PLC), and / or a switch array. The control unit is configured to communicate with one or more external devices including, for example, one or more mechanical pumps, incubators, and / or table shakers. The control unit serves to control at least a portion of the operation of these external devices that are operatively connected to the control unit (e.g., plugged in or wirelessly connected via Bluetooth, WiFi, or other suitable wireless connection) to enable communication between the control unit and the external devices. The control unit is generally programmable in a manner that allows an operator to input one or more sets of instructions into the control unit for operating the system including the tissue processing container, for example, settings for the speed of external devices, such as timing to turn external devices on and off, flow rate, vibration speed, temperature, and / or other variables.

[0046] The term "adjacent," as used herein, may be used to refer to elements that are physically in contact with or next to each other, for example, separated by a small space or gap. The term "central," as used herein, means center, and the term "substantially central" means substantially centered. Although the embodiments described herein are described with the term "central," it is contemplated that each described embodiment may also be described with the term "substantially central."

[0047] In this disclosure, the singular forms "a," "an," and "the" include plural referents unless the context indicates otherwise. Grammatical conjunctions are intended to represent any and all disjunctive and conjunctive combinations of joined clauses, sentences, words, and the like, unless otherwise stated or apparent from the content. Thus, the term "or" should be understood to generally mean "and / or," etc. The term "exemplary" is used in the sense of "example," rather than "ideal." The terms "comprises," "comprising," "includes," "including," or other variations thereof are intended to cover a non-exclusive inclusion, such that a process, method, or product that includes a list of elements does not necessarily include only those elements, but may include other elements not expressly listed or inherent in such process, method, article, or apparatus. In this disclosure, unless otherwise stated, relative terms such as "about," "substantially," and "approximately" are used to indicate that a variation of + / - 10% of the stated value is possible. Moreover, in the claims, values, limits, and / or ranges of the various claimed elements and / or features mean + / - 10% of the stated value, limit, and / or range. In addition, the term "between" when describing a range of values ​​is intended to include the minimum and maximum values ​​set forth herein.

[0048] Tissue Processing Container Tissue processing vessels are provided that are designed and adapted to enhance the efficiency of tissue processing and minimize or reduce undesirable effects on the tissue itself during the processing (e.g., through overly vigorous agitation, abrasions resulting from contact with the surrounding surfaces, damage due to too vigorous flow of processing medium, damage due to physical manipulation or excessive manipulation or pressure placed on the tissue, etc.) The tissue may be of human or animal origin, obtained naturally therefrom, or grown in the laboratory.

[0049] The various processing steps accommodated by such tissue processing containers may generally include, but are not limited to, washing, rinsing, soaking, or otherwise exposing the tissue to a liquid tissue processing medium, such as one or more enzymes, detergents, salt solutions, and / or other desired liquid media, and / or one or more gases suitable for use in tissue processing. The tissue processing container and / or tissue processing medium may further be incubated or brought to a desired temperature. The tissue processing container may also be used to expose one or more tissue specimens to the tissue processing medium and / or one or more gases with agitation, such as by use of a shaker. The tissue processing containers described herein are multifunctional in that the containers accommodate different amounts of liquid in various orientations (bottom down, or bottom up, i.e. right side up, upside down, etc.). Furthermore, the tissue processing containers are stackable, so that the tissue processing containers accommodate the processing of one or more tissue specimens per container, and the processing of multiple tissue specimens across a stack of containers, either in parallel or simultaneously, including as described in more detail below.

[0050] Referring now to the example of FIG. 1A, in some embodiments, FIG. 1A shows an example of a tissue processing container 100 that includes several readily identifiable features and elements that distinguish such containers from other modern dish containers, vessels, flasks, or roller bottles, and the like. That is, it is readily apparent that the tissue processing container 100 as shown in FIG. 1A may be oriented in a right-side-up direction, there may be a central hole 106 across the circular bottom 101 of the tissue processing container 100, and the top of the tissue processing container 100 may be open. The circular bottom 101 possesses an inner side 102 and an outer side 103. The circular bottom 101, the inner side 102, or the outer side 103 may be substantially circular in some aspects. The central hole 106 functions to allow air and liquid to pass through the tissue processing container 100 in a manner that uses the tissue processing container 100 to process tissues, cells, and grafts during processing steps. The central hole 106 need not be of a circular configuration as depicted in Figure 1A, but may be of another configuration such as an oval, elliptical, or polygonal shape. Furthermore, the circular base 101 and the central hole 106 may be of the same shape or may be of different shapes from each other.

[0051] The tissue processing container 100 may be constructed from a polymer or plastic in some embodiments. In one embodiment, the tissue processing container 100 is constructed from one or more of polycarbonate, polyetherimide, polyphenylsulfone, polystyrene, polysulfone, and polymethylpentene, and the like. In another embodiment, the tissue processing container 100 is constructed from polymethylpentene. The tissue processing container 100 may also be constructed from glass or silicate. It will be appreciated that the tissue processing container 100 may be constructed from any material within the scope of the present disclosure.

[0052] The tissue processing container 100 also possesses a first wall 104 possessing a first height surrounding the container such that the container may hold a first volume of liquid. The first volume of liquid held in the container may be between about 100 mL and about 500 mL. In one embodiment, the first volume may be between about 150 mL and about 450 mL. In another embodiment, the first volume may be between about 200 mL and about 400 mL, or between about 250 mL and about 350 mL, or between about 275 mL and about 300 mL. In a specific embodiment, the first volume is about 275 mL. It will be appreciated that the size of the tissue processing container and its features may be increased or decreased to provide for the accommodation of volumes of liquid greater or less than those defined above.

[0053] The area above the tissue processing container 100 as depicted in the orientation shown in FIG. 1A may be open. In other words, there may be no lid on the top of the tissue processing container 100 as depicted in FIG. 1A. In one embodiment, the lid may be seated on top of the first wall 104 such that the interior side 102 is not open, but instead is sealed by the lid. In one embodiment, the optional lid may possess substantially the same diameter as the tissue processing container 100 such that when placed on top of the tissue processing container 100, a seal may be formed that is substantially permeable to air and / or liquid.

[0054] The first wall 104 of the tissue processing vessel 100, in some embodiments, is perpendicular or nearly perpendicular to the circular bottom 101. In other embodiments, the first wall 104 may slope or flare outward from the bottom to the top of the tissue processing vessel 100. That is, in some embodiments, the diameter defined by the top edge of the first wall 104 may be larger at the top than the diameter of the first wall 104 at the bottom of the tissue processing vessel 100 where the first wall 104 is attached to the circular bottom 101. In another embodiment, the first wall 104 may extend upward and outward from where the first wall 104 is attached to the circular bottom 101 so as to form a bowl shape when viewed from the side with rounded walls.

[0055] In one embodiment, the tissue processing vessel 100 may also possess one or more fill reference lines 112, as the tissue processing vessel 100 may be intended to contain a liquid medium at a particular time. The fill reference lines may be horizontal and / or vertical. In one embodiment, there may be a single fill reference line 112, while in other embodiments, there may be multiple fill reference lines 112 indicating above, below, and exactly at a target fill criteria. The one or more fill reference lines 112 may be provided in or on the first wall 104 of any orientation, size, or shape, so long as an operator or scanning instrument can easily determine whether the tissue processing vessel 100 is properly filled with liquid medium during processing, e.g., during a filling step, exposing tissue to tissue processing medium, etc. If the tissue processing container does not contain the desired amount of tissue processing medium, it may indicate that the container was not filled properly, the stack was not assembled properly, there is a problem with the connection to the fill line, a substantially watertight seal was not properly maintained within the stack of tissue processing containers, a broken seal or gasket, or other problem exists, and adjustments may be made to allow for the desired processing of the tissue.

[0056] In some embodiments, the central hole 106 as depicted in Figure 1A may be defined by a second wall 107 having a second height that is less than the first height of the first wall 104. The relative heights of the various features described herein are depicted in the side view of Figure 1B.

[0057] A first mesh screen 108 on the interior side 102 of the tissue processing container is attached to the central hole 106 on the side of the tissue processing container 100 that is open, i.e., the side that is open when the tissue processing container 100 is placed in the orientation shown in FIG. 1A. The first mesh screen 108 may be made, for example, from a metal or a plastic polymer or resin or a combination thereof, and optionally extends over the top of the hole to form a cap structure that completely encases the opening of the central hole 106, i.e., extends along at least a portion of the second wall 107. In one embodiment, the first mesh screen 108 may be constructed from a metal. The metal, in one embodiment, may be stainless steel or other alloys that include one or more types of metal. In another embodiment, the first mesh screen 108 may be constructed from a pure metal, such as aluminum. The metal, in some embodiments, may be coated to reduce or minimize the penetration of tissue specimens into the mesh screen. The gauge or thread count of the mesh may be sufficient such that any contact between the mesh and tissue does not result in unacceptable abrasion or damage to the tissue during tissue processing. In another embodiment, the first mesh screen 108 may be made from one or more plastic polymers. In a specific embodiment, the first mesh screen 108 is made from a polyamide or alternatively a polymer selected from one or more of polybutylene terephthalate (PBT), polyethylene terephthalate (PET), polypropylene (PP), polycarbonate (PC), and the like, or mixtures thereof. In a specific embodiment, the first mesh screen 108 is made from polyamide. In a further embodiment, the first mesh screen is comprised of nylon and / or polypropylene mesh.

[0058] In some embodiments, first gasket 109 surrounds and is rigidly attached to first mesh screen 108. First gasket 109 functions to rigidly seal first mesh screen 108 to second wall 107. In some embodiments, first gasket 109 is comprised of an elastomer or other rubbery polymer or a mixture thereof.

[0059] The one or more troughs 105 are arranged on the inner side 102 of the circular bottom 101. In one embodiment, there are at least two troughs 105 arranged on, rigidly attached to, integrated into, or otherwise formed as part of the inner side 102 of the circular bottom 101. The one or more troughs 105 may be defined by one or more walls that rise at an angle that is approximately perpendicular, but not necessarily perpendicular, to the circular bottom 101. As depicted in FIG. 1A, the one or more troughs 105 are semicircular in shape, but in other embodiments, the one or more troughs 105 are not semicircular, but instead are rectangular, square, circular, triangular, or any other geometric shape. In one embodiment, the one or more troughs 105 may be disposed adjacent to the central hole 106 such that the one or more troughs 105 share a common wall, i.e., the second wall 107, with the central hole 106. In another embodiment, the trough(s) 105 may directly contact the first wall 104 such that one or more walls defining the trough(s) 105 are shared with or defined by the first wall 104. In another embodiment, the trough(s) 105 may possess their own individual walls that do not touch, extend, or coincide in any manner with either the second wall 107 or the first wall 104. In one embodiment, the tissue processing vessel 100 possesses only one trough attached to the interior side 102 of the circular bottom 101. As used herein to discuss one or more troughs, attached may include one or more separate features joined together, integrally formed, incorporated, or the like, so it is clear that attached can include all such configurations. In another embodiment, the tissue processing vessel 100 may possess at least two troughs attached to the interior side 102 of the circular bottom 101. In further embodiments, the tissue processing container 100 may possess at least three or four troughs attached to, incorporated into, or otherwise formed as part of the interior side 102 of the circular bottom 101.

[0060] The one or more troughs 105 may each contain one or more tissue specimens 113. Generally, the one or more troughs 105 may also contain a liquid volume of about 1 mL to 20 mL. For example, in one embodiment, each trough 105 may contain a liquid volume of about 2.5 mL to 17.5 mL, or about 5 mL to 15 mL, or about 7.5 mL to 12.5 mL, or about 10 mL to 12.5 mL. It will again be appreciated that the volume of any trough may be more or less than the aforementioned volumes depending on the size and characteristics of the tissue processing vessel, as well as the number of troughs present in the tissue processing vessel. In some embodiments, each trough 105 contained within the tissue processing vessel 100 may be the same shape and size, while in other embodiments, one or more troughs 105 may have different shapes and / or sizes.

[0061] The second mesh screen 110, optionally bonded, is attached to the exterior side 103 of the circular bottom 101 of the tissue processing container 100. In one embodiment, the second mesh screen 110 may be made from, for example, a metal or a plastic polymer or resin. In one embodiment, the second mesh screen 110 may be constructed from a metal. In another embodiment, the metal may be stainless steel or other alloys containing one or more types of metal. In another embodiment, the second mesh screen 110 may be constructed from a pure metal, such as aluminum. The metal may be coated in some embodiments to reduce or minimize penetration of the tissue specimen through the mesh screen. In another embodiment, the second mesh screen 110 may be constructed from one or more plastic polymers. In a specific embodiment, the second mesh screen 110 may be made from polyamide or alternatively a polymer selected from one or more of polybutylene terephthalate (PBT), polyethylene terephthalate (PET), polypropylene (PP), polycarbonate (PC), and the like, or mixtures thereof. In a specific embodiment, the second mesh screen 110 is made from polyamide. In a further embodiment, the first mesh screen is comprised of nylon and polypropylene mesh. In any embodiment, the first and second mesh screens may be comprised of the same or different materials. The gauge or thread count of the mesh should be sufficient such that any contact between the mesh and tissue does not result in unacceptable abrasion or damage to the tissue during tissue processing.

[0062] In some embodiments, the second gasket 111 may surround the second mesh screen 110. The second gasket 111 may function to seal the second mesh screen 110 against the exterior side 103 of the circular bottom 101. The second gasket 111 may be comprised of an elastomer or other rubber polymer or mixtures thereof in some embodiments. In one embodiment, the second gasket 111 may surround the exterior side 103 of the circular bottom 101 and may be in full direct contact with the first wall 104 along its entire inner periphery. In such an embodiment, the second gasket 111 may be attached to the outside of the first wall 104. In another embodiment, the second mesh screen 110 may seat against the exterior side 103 of the circular bottom 101. Thus, in some embodiments, the second mesh screen 110 may possess approximately the same diameter as the circular bottom 101, while in other embodiments, the second mesh screen 110 may possess a larger diameter than the circular bottom 101. In another embodiment, the second gasket 111 may completely surround or contact the entire perimeter and edges of the second mesh screen 110. In all embodiments, the first gasket and the second gasket may be constructed from the same or different materials.

[0063] In alternative embodiments, one or more struts 114 extending downwardly towards the second mesh screen 110 may be attached to the exterior side 103 of the circular bottom 101. The one or more struts 114 may be constructed of the same material as the tissue processing container 100 in some embodiments. The one or more struts 114 may function to ensure that the second mesh screen 110 does not touch or come into contact with the central hole 106 on the exterior side 103. In some embodiments, the one or more struts 114 may be circular in shape with a diameter smaller than the diameter of the central hole 106. In one embodiment, the tissue processing container 100 may possess at least two struts 114. In another embodiment, the tissue processing container 100 may possess at least three struts 114. In further embodiments, the tissue processing container 100 may possess at least four struts 114. The one or more struts 114 may not necessarily be perpendicular to the circular bottom 101, but in some embodiments, they may be. In other embodiments, the one or more struts 114 may angle inwardly toward the first wall 104 or toward the circular central hole 106. The one or more struts 114 may, in some embodiments, be long enough to extend just beyond the bottom of the tissue processing container 100, and in other embodiments, are only long enough to touch the surface on which the tissue processing container 100 rests in FIG. 1A.

[0064] In one embodiment, the circular bottom 101 may not be flat or perfectly horizontal. That is, in some embodiments, the circular bottom 101 may possess a pitched or inverted cone shape such that the height of the circular bottom 101 at the central circular hole 106 may be higher than the height of the circular bottom 101 at the outer edge or periphery of the circular bottom 101. (See FIG. 1B). That is, in this embodiment, the outer side of the circular bottom 101 may be sloped from the outer diameter to the inner diameter towards the central hole 106 in the center such that when the tissue processing container 100 is oriented in the opposite direction to that depicted in FIG. 1A, i.e., upside down orientation, or when such a circular bottom 101 is flipped 180 degrees in the Y-plane below one or more troughs 105, liquid contacting the outer side will drain through the central hole.

[0065] In further embodiments of the tissue processing container 100, there is a third wall that bisects the circular bottom 101 along approximately the diameter of the circular bottom 101 on the interior side 102. In such embodiments, the third wall may not extend through the circular central hole 106, but instead may only extend outward from the central hole 106 along the interior side 102 of the circular bottom 101 to the first wall 104. In such embodiments, the third wall may have a second height, i.e., a height less than or equal to the height of the second wall 107, and may form or function as one of the walls defining the one or more troughs 105.

[0066] In further embodiments, tissue processing container 100 may include an external second container. The purpose of the second container may be, for example, a graspable shape and / or a platform for a gripping handle or robot for use in automated processing, or other functionality such as stacking stability, color coding, labeling, or otherwise aiding in quick identification of a particular tissue processing container. In this manner, the second container may be of any shape, for example, circular, oval, elliptical or the like, or polygonal, and may be the same or a different shape compared to inner container 100. The second container optionally has a height approximately equal to the height of the first wall. Optionally, the second container is made of the same material as inner container 100. In another embodiment, the second container may be of a height greater or less than the height of the first wall.

[0067] Such tissue processing containers may be beneficial in that one or more troughs 105 may be suitable for containing one or more tissue specimens 113 . Stacked tissue processing containers Tissue processing vessels of the type described herein may be designed to be stacked one on top of the other to form a stack 200. (See FIG. 2A, shown in reverse orientation). When stacking tissue processing vessels, a second mesh screen 210 and / or a second gasket 211 may be placed on top of the tissue processing vessel and another tissue processing vessel 100, which may be oriented in the position shown in FIG. 1A, may be placed on top of the second mesh screen 210 and / or second gasket 211. This process may be repeated until a desired size stack of tissue processing vessels is formed with the second mesh screen 210, the second gasket 211, and the tissue processing vessel 100. Once stacking of tissue processing vessels is completed, the stack may be reversed as shown in FIG. 2A. In the inverted stack 200, the central holes 206 of each tissue processing vessel 100 are aligned linearly with one another as depicted in FIG. 2A.

[0068] As previously mentioned, in one embodiment, the outer diameter of the first wall 204 of the tissue processing container 100 may be larger at the top than at the bottom, thereby creating a top-to-bottom flaring of the first wall 204 outwardly from the central hole 206. In this embodiment, the flaring of the first wall 204 facilitates stacking such that the bottom of a container placed on top of the bottommost tissue processing container 100 fits snugly within the top of the bottommost tissue processing container, and so on throughout the stack 200.

[0069] In some embodiments, the second gasket 211 may be capable of forming a substantially air-tight and substantially water-tight seal between two adjacent tissue processing containers 100 when the adjacent tissue processing containers are stacked and compressed (e.g., pressure is applied to compress the adjacent tissue processing containers together). The second gasket 211 may in some embodiments be capable of a substantially air-tight and substantially water-tight seal by pressure applied between the circular bottom 201 of the tissue processing container 100 and the top edge of the first wall 204 of the upper tissue processing container 100 as oriented in FIG. 2A. In another embodiment, as described above, the second gasket 204 may fit snugly against the surrounding circular bottom 201 and may only touch the inner surface of the first wall 204 of the tissue processing container 100 below when the stack 200 is first formed, thereby forming a substantially water-tight and / or air-tight seal between the inner surface of the first wall 204 of one tissue processing container and the outer edge of the circular bottom 201 of another tissue processing container 100, with the second gasket 211 sandwiched therebetween.

[0070] Referring also to the example of FIG. 2A, in some embodiments, a stack of tissue processing vessels may also include a cap vessel 215 and a base vessel 214 of substantially the same internal dimensions to facilitate stacking with a stack of tissue processing vessels. For example, each tissue processing vessel of a stack of tissue processing vessels as shown in FIG. 2A is inverted (e.g., inverse to FIG. 1B). As depicted in FIG. 2A, each of the cap vessel 215 and the base vessel 214 may possess at least one nozzle 216 and 217, respectively, that traverses through the outer wall of the cap vessel 215 and the base vessel 214, respectively. The nozzle creates a tubular channel from the outside of the cap vessel 215 and the base vessel 214 to the interior space of the cap vessel 215 and the base vessel 214, which may allow a liquid medium or gas to pass therethrough and into the adjacent tissue processing vessel through the central hole of that tissue processing vessel. In some embodiments, each of the cap container 215 and the base container 214 may include an open interior space through which liquid and / or gas may be transported through a nozzle and a hose connected to an external liquid / gas source that is then transported into the open space of the bottom (formerly cap) container and then through their central holes to each of the tissue processing containers in the stack. In such embodiments, a tube or hose directly or indirectly connected to a source of liquid (or gas, as the case may be, including but not limited to tissue processing medium) may in some embodiments be rigidly attached to the nozzle on the outside of the stack 200, such that when liquid, for example, flows therethrough, the liquid is directed from the outside of the stack through the nozzle to the inside of the stack and, in the case of the base container 214, up through the central hole 206 of the bottom-most tissue processing container 100 in the stack 200. Conversely, the nozzle 216 of the cap vessel may similarly traverse the outer wall of the cap vessel 215 , advance into a tube within the cap vessel 215 , and advance to a position directly above the central hole 206 in the center of the topmost tissue processing vessel 100 in the stack 200 .In this manner, for example, liquid directed into the topmost container 100 may travel through the central hole 206 in the center of the topmost container, then through a tube in the cap container 215, and then exit the stack 200 via the nozzle 216. In some embodiments, the cap container 215 and base container 214 may be optionally added to the stacked tissue processing containers 200, after which the stack 200 may be tilted to the orientation shown in Figure 2A, i.e., the cap container 215 and base container 214 may be inverted in the stack of tissue processing containers before or after being added to the stack.

[0071] An alternative embodiment of the cap vessel 215 and base vessel 214 is provided in Figure 2B. It can be readily inferred from the combined cap / base vessel depicted in Figure 2B that the same cap / base element is used for both the top and bottom of the stack.

[0072] When tissue processing containers 100 are stacked on top of each other in this manner to form a stack 200, and then inverted and a cap container 215 and a base container 214 are added to the top and bottom of the stack 200, respectively, a substantially air-tight and substantially water-tight seal is formed due to pressure exerted on the second mesh screen 210 and second gasket 211 interposed between each tissue processing container 100, and may be formed directly between the top-most and bottom-most tissue processing containers and the cap / base container. Furthermore, in such a configuration, liquid dispensed through nozzle 217 in base container 214 fills the bottom-most tissue processing container 100 first through central hole 206. The liquid level gradually rises in the bottom-most container and, in one embodiment, first contacts the top-most edge of second wall 207 surrounding central hole 206.

[0073] As noted above, in one embodiment, the circular bottom 201 may not be flat. In this embodiment, the top-most edge of the second wall 207 in the orientation depicted in FIG. 2A would be the first interior side element contacted by the liquid as it rises inside the tissue processing container 100. In doing so, an air pocket is formed at the top of the tissue processing container 100, above the final liquid level within the tissue processing container 100, so that the interior side 202 of the circular bottom 201 does not touch the liquid dispensed through the circular central hole 206. In this embodiment, the air pocket thus formed may serve the purpose of enhancing or maximizing the mixing of the liquid and exposing the tissue to the liquid during various tissue processing steps (such as when the tissue processing container stack 200 is shaken by use of a shaker, as described in more detail below).

[0074] Furthermore, when two or more tissue processing containers 100 are stacked on top of each other, if liquid and / or tissue 213 is present in one or more troughs 205, when the stack 200 is inverted, the liquid and / or tissue 213 present in the one or more troughs 105 will come into contact with the second mesh screen 210. The liquid that was present in the one or more troughs 205 will then pass through the second mesh screen 210 and through the circular central hole 206 of the tissue processing container 100 below. This flow of liquid previously contained by the one or more troughs 205 continues through the stack until it reaches the nozzle 217 and exits the stack 200. Meanwhile, the tissue specimens 213 in each tissue processing container 100 rest on the mesh screen 210 below.

[0075] Thus, in one embodiment, tissue processing container 100 may be constructed from a material that does not pierce, bind, or adhere to tissue 113 or 213. Additionally, first mesh screen 208 and second mesh screen 210 may be constructed from a material that does not pierce, bind, or adhere to tissue 113 or 213. In certain embodiments, one or more components of tissue processing container 100 may be further coated with one or more additional chemicals, polymers, biological materials, buffering materials, or other substances that reduce or minimize adverse effects on tissue when in contact with such components of the tissue processing container. In a specific embodiment, tissue processing container 100 is made from glass.

[0076] In another embodiment, each of the first walls 204 of the tissue processing containers 100 further possesses a lip or skirt extension material that overlaps the periphery of the circular bottom 201 of the tissue processing container 100 below when oriented as shown in FIG. 2A such that a substantially air-tight and substantially water-tight seal is formed between each tissue processing container 100 in the stack 200 by a second gasket 211 that fits tightly or snugly between the exterior surface of the first wall 204 of one tissue processing container 100 on the bottom and the interior surface of the first wall 204 of another tissue processing container 100 resting thereon in the stack 200. An additional function that said skirt or lip material of the first walls 204 may serve is to increase the stability of the stack 200. The extended material of the first wall 204 described above, which extends across the circular bottom 201 of each tissue processing container 100 in the stack 200, may provide additional mechanical stability to the entire stack 200, which may at times help maintain a substantially air-tight and substantially water-tight seal between each tissue processing container 100 during processing steps such as tilting or inverting the stack, moving the stack, and / or agitating the stack 200 with a shaker, and the like.

[0077] Systems Including Tissue Processing Containers - Patent application Further contemplated herein is a system 300 for processing tissue, including a tissue processing container 100, for example as shown in Figures 3A and 3B and as otherwise described herein. The contemplated system includes various components, including but not limited to, two or more tissue processing containers 100 forming one or more stacks 200, optionally each stack 200 further including a cap container 315 and a base container 314 as shown in Figure 1B, and a transport housing 301 encasing the one or more stacks 200. The transport housing 301 is of sufficient dimensions to encase the one or more stacks 200. The transport housing 301 may provide protection for safely transporting the one or more stacks 200 from a first location to a second location, optionally during desired processing, and thus may act as a carrying case for the stacked tissue processing containers. In some embodiments, the transport housing also includes a stabilizer shroud for each stack of tissue processing containers it houses. Such stabilizer shrouds are used to maintain the stability of each stack of tissue processing containers and to ensure that each fits snugly within the transport housing to avoid accidental disassembly of the stacks and to avoid breaking the substantially air-tight and substantially water-tight seal between the cap and base containers of each stack of tissue processing containers. The transport housing may further include a cover for enclosing and / or sealing the one or more stacks of tissue processing containers.

[0078] In some embodiments, the transport housing 301 fits inside the incubator 302. In other embodiments, one or more stacks 200 are placed directly into the incubator 302 and the transport housing 301 is not needed, for example depending on the processing step and the tissue being processed. In another embodiment, the transport housing 301 optionally includes an air regulator to help control and maintain the appropriate gases required for any particular stage of tissue processing. Such gas components include, but are not limited to, oxygen, nitrogen, carbon dioxide, and / or argon or other inert gases, as well as moisture (water) in the air to maintain the appropriate humidity level of the tissue during processing. In another embodiment, the incubator 302 includes and is equipped with such gas and humidity control devices.

[0079] In one embodiment, the incubator 302 may be placed on top of a vibrating shaker fixture 306. The vibrating shaker is not particularly limited and many are commercially available. However, in certain embodiments, the incubator 302, transport housing 301, and shaker 306 may all be controllable by a central processing unit or controller 305. In another embodiment, one or more stacks of tissue processing containers 200 may be placed directly on top of the shaker 306 inside the incubator 302. Any number of such variations in system configuration are possible, so long as the tissue inside the tissue processing container 100 is exposed to the appropriate atmospheric conditions, temperature, and mixing necessary for the particular processing of the tissue.

[0080] It should be noted that to achieve complete exposure of the tissue to the tissue processing medium, the tissue processing container 100 may be vibrated or shaken at a rate that achieves this goal. Surprisingly, it has been discovered that by designing the tissue processing container 100 as described herein, including an air pocket above the liquid level line, and by vibrating or shaking the container at a desired frequency, a standing wave of mixing can be established within the tissue processing container such that complete and reproducible processing of the biological material is achieved. Optionally, the shaker 306 may include a tilting or tiltable function in addition to the rocking / vibrating function that is standard on commercially available laboratory shakers. Further optionally, the tissue processing stack 200 may also be tilted at a specific frequency to achieve complete mixing and exposure of the tissue to the liquid medium.

[0081] In some embodiments, as shown in FIG. 3A, the system may also include various tubes / hoses 303 that provide for the ingress of fluids and egress of waste and air to a waste container 307. The hoses may be attached to nozzles 317 and 316 to provide channels to direct liquids into and out of the tissue processing container 100. One or more electric / mechanical solution pumps 304 are also provided by the systems described herein. The solution pumps may function to generate pressure within the hoses 303, thereby effecting the ingress and egress of fluid media solutions into and out of the tissue processing stack 200 via the nozzles 317 and 316. The pumps 304 are optionally controllable and may be linked to a controller unit 305, as well as other hardware elements described above.

[0082] In some embodiments of the described systems, one or more media reservoirs 301 containing one or more tissue processing media are further provided for use in processing (e.g., washing, rinsing, soaking, culturing, etc.) the tissue specimens in the tissue processing container 100.

[0083] Additionally, one or more programmable controller units 305 may be provided that can be linked electrically or by electrical signals, such as Wi-Fi, Internet, etc., to the various hardware elements described above, i.e., incubator 302, one or more pumps 304, shaker 306, housing 301, and the like. The linking allows the programmable controller unit 305 to automate processing protocols for each biological tissue, cell, or graft in the tissue processing vessel. In some embodiments, the programmable controller unit may provide a user interface, programmable memory, computing chips, and other computer components necessary to run software programs that control not only the start / stop functions of each hardware component, but also other variables specific to each hardware component, such as temperature, vibration frequency, pitch or tilt, pump speed, and the like. In this manner, the described system may allow an operator to freely program one or more, e.g., all, of the variables into the programmable controller unit for processing tissue specimens in the tissue processing vessel. Such automated features may provide a variety of benefits, including saving time and materials, improving tissue processing results, uniformity and reproducibility of processing, and other efficiencies.

[0084] How to use the tissue processing container The tissue processing containers described herein may be useful for different tissue processing protocols. The systems described above, e.g., automated systems, may be customizable to meet the needs of any desired tissue processing step. To further describe such methods, non-limiting exemplary embodiments are provided below for illustrative purposes.

[0085] In one embodiment, the tissue processing container described herein may be utilized for processing neural tissue. In such a method, an empty tissue processing container as described herein may be oriented such that the interior side of the bottom faces upward. Optionally, the tissue processing container may be placed on top of a base container. A debrided neural tissue specimen may be placed in each trough of the tissue processing container, and a first tissue processing medium may also be added to each trough containing a tissue specimen. Alternatively, the first tissue processing medium may be added to the trough before or at the same time that a tissue specimen may be placed in the same trough. In one embodiment, the first tissue processing medium may include one or more active enzymes.

[0086] The first liquid medium may be added manually, for example using a calibrated pipette. Alternatively, one or more additions of the tissue specimen and the first tissue processing medium may be automated, for example through a machine programmed to deliver the tissue specimen and / or tissue processing medium to the desired trough in a reproducible manner.

[0087] Any portion of these steps may be repeated with the addition of tissue processing vessels and tissue specimens as desired to provide a desired number of tissue processing vessels containing tissue specimens and tissue processing medium. Additional tissue processing vessels may optionally be stacked. The stacking may occur such that the additional processing vessels are stacked on top of each other after each receives a tissue specimen and tissue processing medium, or a tissue processing vessel may be added to the stack before the tissue processing vessel receives one or more of the tissue specimens and tissue processing medium, a tissue specimen and / or tissue processing medium may then be added to it, another tissue processing vessel may be added thereon, and so on until a desired stack size is achieved.

[0088] In some embodiments, as stacking of tissue processing vessels proceeds, cap vessels and base vessels may be further added to the stack. Screens and gaskets may be included in the stack between each vessel comprising the stack. The above steps may be repeated until a desired number of stacks is achieved, and then one or more stacks (e.g., three stacks) may be placed into and sealed in a mobile housing. The mobile housing containing the stack of tissue processing vessels may be placed inside an incubator at a desired first temperature for a first period of time, either manually or, for example, via a robotic arm (or the mobile housing may itself be used to incubate the vessels, if so equipped). For example, if the first tissue processing medium is an enzyme, the mobile housing containing the stack of tissue processing vessels may be incubated at an optimal incubation temperature for that particular enzyme. Such temperatures and periods of incubation with each are readily known or discernible by one of skill in the art for each enzyme that may be used. This same incubation step may be used for tissue processing vessels that are not stacked and / or that are stacked but not placed inside a mobile housing.

[0089] Once the first period is complete, if the tissue processing containers have not yet been stacked, each may be stacked on top of the other as described above until the desired stack size is achieved. In some embodiments, a base container and a cap container are added to the stack of tissue processing containers, although as noted above, the base container and cap container may be added during the formation of the stack as part of the stack. In one embodiment, the tissue processing container may already include both the first and second mesh screens, and the first and second gaskets, prior to stacking. In other embodiments, the tissue processing container may only include the first mesh screen and the first gasket, and the second mesh screen and second gasket may be added immediately prior to stacking the tissue processing containers, by adding the tissue processing container, the second gasket, and then the second mesh screen (this order may be reversed in some embodiments), followed by the second tissue processing container. This process of adding these three components may then be repeated until the desired number of stacks is achieved.

[0090] After the stacked tissue processing containers have completed the first incubation period, the transport housing (or individual stacks) may be inverted to allow the first tissue processing medium to be drained away from the tissue specimens and exit the tissue processing containers. The first tissue processing medium may then exit through a nozzle in the cap container (which is now at the bottom of the stack) and into one or more waste receptacles.

[0091] In some embodiments, the stacked tissue processing containers and / or transport housings may then be returned to the incubator, or depending on the tissue processing protocol, the next stage of the tissue process may be performed without the use of an incubator. If the next step proceeds using an incubator, the temperature of the incubator may be adjusted to a second desired temperature for the next culture period before or after the processing containers and / or transport housings containing the processing containers are returned to the incubator. In some embodiments, the cap container and base container may be attached to appropriate tubing / hoses to automatically or manually add and possibly remove a second tissue processing medium and / or liquid and / or gas. When the desired temperature is reached, the second liquid medium may be added manually or automatically via a pre-programmed mechanism through a nozzle in each cap container (e.g., as shown in FIG. 3A where the stacked tissue processing containers are inverted) into the stacked tissue processing containers, and thus into each of the tissue processing containers in the stack. As described above, the tissue processing medium travels through the hose, through the nozzle of the cap container, into the hole of the bottommost tissue processing container, and fills the bottommost tissue processing container until the liquid level line reaches the tip or edge of the second wall of the central hole. The liquid then enters the central hole of the tissue processing container above the bottommost tissue processing container and proceeds into the second tissue processing container of the stack, and so on until each of the stacks of tissue processing containers contains an appropriate amount of tissue processing medium. This process may result in air pockets above the liquid level line of one or more or each of the tissue processing containers, enhancing optimal mixing and agitation of the liquid medium.

[0092] At each stage, after all the tissue processing containers are filled, a shaker may optionally be used to vibrate the tissue processing containers at a desired frequency. In one embodiment, to enhance mixing or achieve maximum mixing, the revolutions per minute (rpm) of the shaker may be adjusted until a standing wave is achieved within the tissue processing container.

[0093] In one embodiment, before proceeding with processing, the fill criteria of each tissue processing vessel is inspected to ensure that an appropriate amount of second tissue processing medium is present in each of the tissue processing vessels. In one embodiment, the fill criteria is determined by visually inspecting each tissue processing vessel. In another embodiment, the fill criteria is determined by visually inspecting a fill line optionally placed on the first wall of each tissue processing vessel. In a further embodiment, the evaluation of the fill criteria of each tissue processing vessel may be performed by a camera connected to a programmable controller unit, and a software program is optionally stored that identifies which tissue processing vessels have been properly filled and which have not, based on the data provided through the camera. Such inspection may ensure that there are no leaks between the tissue processing vessels and that the settings have been properly achieved.

[0094] The tissue processing vessels containing the second tissue processing medium may then be incubated at a desired second temperature for a second period of time, after which the second liquid medium may be vented from one or more, such as all, of the tissue processing vessels by opening a nozzle in a cap vessel (e.g., as shown in FIG. 3A where the stacked tissue processing vessels are inverted), and the second liquid medium can be drained into one or more waste vessels.

[0095] This process of filling and draining the liquid medium may be repeatable as many times as desired. The filling and draining steps may be controllable by the programmable controller unit described above. Similarly, the programmable controller unit, in some embodiments, is pre-programmed to control one or more external hardware devices in the system, such as one or more shakers, incubators, one or more pumps, etc. Thus, in some embodiments, the above method may be fully automated in some embodiments by the programmable controller unit, or may be manual in other embodiments.

[0096] In one embodiment, filling and emptying the stacked tissue processing containers may include the use of a tissue processing medium to wash and / or rinse the tissue specimens contained within the tissue processing containers. In some embodiments, one or more steps may be repeated and / or the tissue processing medium or media utilized in each filling step may be the same or different in terms of ingredients or composition. Each step of filling and emptying the tissue processing containers in this manner may both be automated and may be customizable to a desired protocol for processing and for the particular type of tissue to be processed.

[0097] In one embodiment, the first period of time may be different from the second period of time. In another embodiment, the first period of time and the second period of time are the same. In one embodiment, the first temperature is the same as the second temperature. In a further embodiment, the first temperature and the second temperature are different.

[0098] After processing the tissue, the stacked tissue processing containers may be removed from the incubator and / or housing and separated from the container base and cap. The stack may then be separated to provide access to the processed tissue, which rests on top of the second mesh screen at the bottom of each of the tissue processing containers for further use.

[0099] Further modifications and alternative embodiments of various aspects of the methods and systems described herein will be apparent to those skilled in the art upon consideration of this specification. This specification should therefore be construed as illustrative only, and is intended to teach those skilled in the art the general manner of carrying out the disclosed methods and systems. It should be understood that the forms of the disclosed methods and systems shown and described herein should be taken as examples of embodiments. Elements and materials may be substituted for those illustrated and described herein, parts and steps may be reversed, and certain features of the disclosed methods and systems may be utilized independently, all as would become apparent to those skilled in the art after having the benefit of this description of the disclosed methods and systems. Changes may be made to the elements described herein without departing from the spirit and scope of the disclosed methods and systems, as set forth in the following claims.

[0100] The scope and breadth of the present disclosure should not be limited by any of the above exemplary embodiments, but should be defined only by the following claims and their equivalents. That is, the above examples are included to demonstrate various exemplary embodiments of the described methods and systems. Those skilled in the art will recognize that the techniques disclosed in the examples represent techniques discovered by the inventors to work well in the practice of the described methods and systems, and thus may be considered to constitute optional or exemplary modes for the practice thereof. However, those skilled in the art will recognize in light of the present disclosure that many changes can be made to these specific embodiments disclosed and still obtain the same or similar results without departing from the spirit and scope of the described methods and systems. The technical ideas contained in the present disclosure are described below. (Supplementary Note 1) A container system comprising: a cup having an outer wall; an inner wall disposed radially inward of the outer wall, the inner wall having a height less than a height of the outer wall; a floor of the cup having a hole substantially in a center of the floor, the hole being disposed radially inward of the inner wall such that the inner wall surrounds the hole and the outer wall surrounds the inner wall; and at least two troughs disposed on an inner side of the floor of the cup, the at least two troughs surrounding the hole and the at least two troughs being defined by two or more trough walls protruding from the inner side of the floor. (Appendix 2) The container system of Appendix 1, further comprising a mesh screen covering the hole. (Appendix 3) The container system of Appendix 2, further comprising a gasket configured to tightly seal the mesh screen against the inner wall. (Appendix 4) The container system of Appendix 1, further comprising a mesh screen disposed on an exterior side of the floor opposite the interior side. (Appendix 5) The container system of Appendix 4, further comprising a gasket configured to seal the mesh screen against the exterior side of the floor or the exterior side of the exterior wall. (Appendix 6) The container system of Appendix 1, wherein the outer wall extends radially outward such that the outer wall extends away from the floor of the cup. (Appendix 7) The container system of Appendix 1, wherein the floor is sloped so that the floor extends from the outer wall to the inner wall. (Additional Note 8) The container system of Additional Note 1, wherein the end of the cup opposite the floor is not covered. (Appendix 9) The container system of appendix 1, wherein the substantial center of the floor is the center of the floor. (Supplementary Note 10) A container system comprising: a cup having a periphery defined by an outer wall, the cup having a closed end and an open end; an inner wall disposed radially inward of the outer wall, the inner wall having a height less than a height of the outer wall; a floor defining the closed end of the cup; a hole disposed substantially in a center of the floor, the hole disposed radially inward of the inner wall such that the inner wall surrounds the hole and the outer wall surrounds the inner wall; at least two troughs disposed on an inner surface of the floor of the cup, the at least two troughs being defined by two or more trough walls protruding from the floor toward the open end of the cup, the two or more trough walls each having a height less than the height of the inner wall; a first mesh screen extending across the hole; and a second mesh screen extending along an outer surface of the closed end of the cup. (Appendix 11) The container system of Appendix 10, further comprising a first gasket configured to tightly seal the first mesh screen across the hole, and a second gasket configured to seal the second mesh screen to the closed end of the cup. (Appendix 12) The container system of Appendix 11, wherein an end of the outer wall closest to the floor of the cup defines a first diameter, an end of the inner wall closest to the floor of the cup defines a second diameter smaller than the first diameter, and the hole defines a third diameter smaller than the second diameter. (Addendum 13) The container system of Addendum 12, wherein the floor of the cup slopes radially outward from the second diameter to the first diameter such that the floor adjacent the first diameter is positioned farther from the open end of the cup than the floor adjacent the second diameter. (Supplementary Note 14) The container system of Supplementary Note 10, further comprising a lid configured to removably cover the open end of the cup. (Supplementary Note 15) The container system of Supplementary Note 10, wherein the substantial center of the floor is the center of the floor. (Supplementary Note 16) A container system, comprising a first cup and a second cup, each of the first cup and the second cup having an open end and a closed end, an outer wall extending from the open end to the closed end and defining a periphery, an inner wall disposed radially inward of the outer wall, the inner wall having a height less than the height of the outer wall, and a floor having a hole substantially in the center of the floor, the hole being disposed radially inward of the inner wall such that the inner wall surrounds the hole and the outer wall surrounds the inner wall, and a floor on an inner surface of the floor of the cup. and at least two troughs disposed in a space between the floor and the container, the at least two troughs being defined by two or more trough walls protruding from the interior surface of the floor, the closed end of the first cup being dimensioned to be received within the open end of the second cup such that the first cup is stackable with the second cup, and the hole in the first cup aligns with the hole in the second cup when the first cup is stacked with the second cup. (Additional Note 17) The container system of Additional Note 16, wherein each of the first cup and the second cup further includes a first mesh screen extending across the hole. (Additional Note 18) The container system of Additional Note 17, wherein each of the first cup and the second cup further includes a gasket that securely secures the first mesh screen across the hole. (Additional Note 19) The container system of Additional Note 18, wherein each of the first cup and the second cup further includes a second mesh screen extending along an exterior surface of the closed end of the cup. (Supplementary Note 20) The container system of Supplementary Note 19, wherein each of the first cup and the second cup further includes a second gasket securing the second mesh screen along the exterior surface of the closed end of the cup. (Addendum 21) The container system of Addendum 16, wherein the outer wall of the first cup and the outer wall of the second cup each define a first diameter adjacent the closed end that is smaller than a second diameter adjacent the open end. (Addendum 22) The container system of Addendum 16, further comprising: a base container having a first nozzle extending into an interior region of the base container, the first nozzle configured to be fluidly connected to the hole of the first cup; and a cap container having a second nozzle extending into an interior region of the cap container, the second nozzle configured to be fluidly connected to the hole of the second cup. (Additional Note 23) The container system of Additional Note 16, wherein the substantial center of the floor is the center of the floor. (Appendix 24) 21. The container system of any one of Appendices 10 to 15, 19, and 20, wherein the gauge of the first mesh screen and / or the second mesh screen is determined so that contact between the first mesh screen and / or the second mesh screen and tissue does not cause abrasion or damage to tissue during tissue processing. (Appendix 25) 25. The container system of any one of claims 10-15, 19, 20, and 24, wherein the gauge of the first mesh screen and / or the second mesh screen is from about 300X300 threads per inch to about 500X500 threads per inch. (Appendix 26) 26. The container system of claim 25, wherein the gauge of the first mesh screen and / or the second mesh screen is approximately 325x325 threads per inch. (Appendix 27) 25. The container system of any one of claims 10-15, 19, 20, and 24, wherein the first mesh screen and / or the second mesh screen have openings of less than about 43.18 μm (0.0017 inches). (Appendix 28) 19. The container system of any one of claims 2 to 5, 17, and 18, wherein the mesh screen has a gauge such that contact between the mesh screen and tissue does not cause abrasion or damage to the tissue during tissue processing. (Appendix 29) 29. The container system of any one of claims 2-5, 17, 18, and 28, wherein the mesh screen has a gauge of about 300X300 threads per inch to about 500X500 threads per inch. (Appendix 30) 30. The container system of claim 29, wherein the gauge of the mesh screen is approximately 325x325 threads per inch. (Appendix 31) 29. The container system of any one of claims 2-5, 17, 18, and 28, wherein the mesh screen has openings of less than about 43.18 μm (0.0017 inches). (Appendix 32) 16. The container system of any one of claims 1 to 15, further comprising an air regulator that controls and maintains the appropriate gas required, and a humidity control device that maintains a predetermined humidity standard of the tissue during processing of the tissue in the cup. (Appendix 33) an incubator configured to control at least a temperature of the tissue and tissue processing medium in the cup; a shaker configured to tilt and / or vibrate the cup; a camera configured to assess a fill standard of the tissue processing medium in the cup; one or more pumps connected to the container system via one or more hoses and configured to supply the tissue processing medium to the cup, the one or more pumps configured to generate pressure within the one or more hoses to effect entry and exit of the tissue processing medium into the cup via a nozzle; 33. The container system of any one of claims 1 to 15 and 32, comprising one or more of the following: (Appendix 34) 34. The container system of claim 33, further comprising a controller having a user interface, a programmable memory, and a computing chip, the controller configured to execute a software program that controls one or more of the incubator, the shaker, the camera, and the one or more pumps. (Appendix 35) 24. The container system of any one of claims 16 to 23, further comprising an air regulator that controls and maintains the appropriate gas required, and a humidity control device that maintains a predetermined humidity standard of tissue during processing of the tissue in the first cup and the second cup. (Appendix 36) an incubator configured to control at least a temperature of the tissue and tissue processing medium in the first cup and the second cup; a shaker configured to tilt and / or vibrate the first cup and the second cup; a camera configured to assess a fill standard of tissue processing medium in the first cup and the second cup; one or more pumps connected to the container system via one or more hoses and configured to supply the tissue processing medium to the first cup and the second cup, the one or more pumps configured to generate pressure within the one or more hoses to effect entry and exit of the tissue processing medium into and from the first cup and the second cup via nozzles; 36. The container system of any one of claims 16 to 23 and 35, comprising one or more of the following: (Appendix 37) 37. The container system of claim 36, further comprising a controller having a user interface, a programmable memory, and a computing chip, the controller configured to execute a software program that controls one or more of the incubator, the shaker, the camera, and the one or more pumps. (Appendix 38) 38. The container system of any one of claims 16-23, 35, 36, and 37, further comprising a plurality of cups including the first cup and the second cup, the plurality of cups being arranged in one or more stacks. (Appendix 39) 40. The container system of claim 38, further comprising a housing including a cover and configured to encase the one or more laminates. (Appendix 40) 40. The container system of any one of claims 38 to 39, further comprising a stabilizer shroud for each ply of the one or more plies. (Appendix 41) 41. The container system of any one of claims 1 to 40, wherein the two or more trough walls are perpendicular to an interior side of the floor. (Appendix 42) 42. The container system of any one of claims 1 to 41, wherein each of the at least two troughs is in direct contact with the outer wall such that one or more of the two or more trough walls are shared with or defined by the outer wall. (Appendix 43) 42. The container system of any one of claims 1 to 41, wherein the two or more trough walls do not touch, extend, or coincide in any way with either the inner wall or the outer wall. (Appendix 44) 44. The container system of any one of claims 1 to 43, wherein the floor of the cup is sloped such that when an inner side of the floor is facing upward, the hole in the floor of the cup is higher than an outer edge of the floor of the cup.

Claims

1. 1. A tissue processing container system comprising: A first cup and a second cup, each of the first cup and the second cup comprising: an open end and a closed end; an outer wall extending from the open end to the closed end and defining a perimeter; an inner wall disposed radially inward of the outer wall, the inner wall having a height less than a height of the outer wall; a floor having a hole substantially at a center of the floor, the hole being disposed radially inward of the inner wall such that the inner wall surrounds the hole and the outer wall surrounds the inner wall; at least two troughs disposed on an interior surface of the floor of the cups, the at least two troughs being defined by two or more trough walls projecting from the interior surface of the floor; the closed end of the first cup is sized to be received within the open end of the second cup such that the first cup is stackable with the second cup, and the hole in the first cup is configured to align with the hole in the second cup when the first cup is stacked with the second cup; each of the first cup and the second cup further comprising a first mesh screen, the first mesh screen of the first cup extending across the hole in the first cup and the first mesh screen of the second cup extending across the hole in the second cup.

2. The tissue processing container system of claim 1 , wherein the two or more trough walls each have a height less than the height of the inner wall.

3. 3. The tissue processing container system of claim 1, wherein each of the first cup and the second cup further comprises a first gasket, the first gasket of the first cup securely fixing the first mesh screen across the hole of the first cup, and the first gasket of the second cup securely fixing the first mesh screen across the hole of the second cup.

4. 4. The tissue processing container system of claim 1, wherein each of the first cup and the second cup further comprises a second mesh screen, the second mesh screen of the first cup extending along an exterior surface of the closed end of the first cup and the second mesh screen of the second cup extending along an exterior surface of the closed end of the second cup.

5. 5. The tissue processing container system of claim 4, wherein each of the first cup and the second cup further comprises a second gasket, the second gasket of the first cup securing the second mesh screen of the first cup along the exterior surface of the closed end of the first cup, and the second gasket of the second cup securing the second mesh screen of the second cup along the exterior surface of the closed end of the second cup.

6. 6. The tissue processing container system of claim 1, wherein the outer wall of the first cup and the outer wall of the second cup each define a first diameter adjacent the closed end that is smaller than a second diameter adjacent the open end.

7. a base container having a first nozzle extending into an interior region of the base container, the first nozzle configured to be fluidly connected to the bore of the first cup; 7. The tissue processing container system of claim 1, further comprising: a cap container having a second nozzle extending into an interior region of the cap container, the second nozzle configured to be fluidly connected to the hole in the second cup.

8. The tissue processing container system of any one of claims 1 to 7, wherein the substantial center of the bed is the center of the bed.

9. 6. The tissue processing container system of claim 4 or 5, wherein the gauge of the first mesh screen and / or the second mesh screen is determined so that contact between each of the first mesh screen and each of the second mesh screen and tissue does not cause abrasion or damage to the tissue during tissue processing.

10. 10. The tissue processing container system of any one of claims 4, 5 and 9, wherein each first mesh screen and / or each second mesh screen has a gauge of about 300x300 threads per inch to about 500x500 threads per inch.

11. 11. The tissue processing container system of claim 10, wherein the gauge of each first mesh screen and / or each second mesh screen is approximately 325x325 threads per inch.

12. 10. The tissue processing container system of any one of claims 4, 5 and 9, wherein each first mesh screen and / or each second mesh screen comprises openings of less than about 43.18 μm (0.0017 inches).

13. 13. The tissue processing container system of claim 1, further comprising an air regulator to control and maintain the appropriate gas required, and a humidity control device to maintain a predetermined humidity standard of tissue during processing of the tissue in the first cup and the second cup.

14. an incubator configured to control at least a temperature of the tissue and tissue processing medium in the first cup and the second cup; a shaker configured to tilt and / or vibrate the first cup and the second cup; a camera configured to assess a fill standard of tissue processing medium in the first cup and the second cup; one or more pumps connected to the tissue processing container system via one or more hoses and configured to supply the tissue processing medium to the first cup and the second cup, the one or more pumps configured to generate pressure within the one or more hoses to effect entry and exit of the tissue processing medium into and from the first cup and the second cup via nozzles; The tissue processing container system according to any one of claims 1 to 13, comprising one or more of the following:

15. 15. The tissue processing container system of claim 14, further comprising a controller having a user interface, a programmable memory, and a computing chip, the controller configured to execute a software program that controls one or more of the incubator, the shaker, the camera, and the one or more pumps.

16. The tissue processing container system of any one of claims 1 to 15, further comprising a plurality of cups including the first cup and the second cup, the plurality of cups being arranged in one or more stacks.

17. 17. The tissue processing container system of claim 16, further comprising a housing including a cover and configured to encase the one or more stacks.

18. 18. The tissue processing container system of claim 16 or 17, further comprising a stabilizer shroud for each stack of the one or more stacks.

19. The tissue processing container system of any one of claims 1 to 18, wherein the two or more trough walls are perpendicular to an interior side of the floor.

20. The tissue processing container system of any one of claims 1 to 19, wherein each of the at least two troughs is in direct contact with the outer wall such that one or more of the two or more trough walls are shared with or defined by the outer wall.

21. The tissue processing container system of any one of claims 1 to 19, wherein said two or more trough walls do not touch, extend or coincide in any manner with either said inner wall or said outer wall.

22. 22. The tissue processing container system of any one of claims 1 to 21, wherein the floor of the cup is sloped such that when the inner side of the floor is facing upwards, the hole in the floor of the cup is higher than the outer edge of the floor of the cup.

Images