Biological Modeling Device

Abstract

The present invention provides a biological modeling device for co-cultivation of cells, the device comprising a first chamber providing a first reservoir and a second chamber providing a second reservoir, the second reservoir and the first reservoir being connected via a permeable support, the first chamber being at least partially received within the second chamber, the device further comprising a seal positioned to separate a first gaseous environment of the first reservoir in the first chamber from a second gaseous environment of the second reservoir in the second chamber and sealably engaging the first and second chambers.

Description

[Technical Field]

[0001] The present invention relates generally to biological modeling devices, and more particularly to a biological modeling device having two chambers, each having a gaseous environment, with an improved sealing mechanism for maintaining separation between the gaseous environments of the two chambers. [Background technology]

[0002] Biological modeling devices are known to be used for in vitro culturing of biological cells and / or tissues to simulate an in vivo environment, such as a luminal model of the digestive tract. Typically, biological modeling devices have a plate with one or more depressions or "wells" on the apical side that are used to receive biological elements, such as cell populations or tissue samples. Some devices also have one or more cup-shaped inserts, each positioned within a well. The inserts receive and hold a different biological element, such as a second cell population or tissue sample, on the basolateral side. Some known inserts have a membrane on the bottom surface. When a nutrient medium is introduced into one or more wells and / or inserts of the plate, the biological element(s) receive nutrients from the medium.

[0003] It would be desirable to provide a biological modeling device with an improved sealing mechanism that maintains separation between the gaseous environments of the two chambers. In particular, it is an object of the present invention to provide a biological modeling device that provides a sealed, separated gaseous environment between the well(s) and the insert(s). It is an object of the present invention to provide a biological modeling device in which the gas transfer path between the well(s) and the insert(s) is substantially through an adjacent membrane.

[0004] The present invention provides at least an alternative to prior art biological modeling devices. Summary of the Invention

[0005] According to the present invention there is provided a biological modelling apparatus according to the appended claims.

[0006] According to one aspect of the present disclosure, there is provided a biological modeling apparatus comprising two chambers: the first chamber provides a first reservoir having a first gaseous environment, the second chamber provides a second reservoir having a second gaseous environment, the second reservoir and the first reservoir are connected via a permeable support, and the first chamber is at least partially received within the second chamber; The device includes a seal disposed to separate a first gaseous environment of the first reservoir from a second gaseous environment of the second reservoir and in sealable engagement with the first and second chambers.

[0007] In certain embodiments, the permeable support is a membrane.

[0008] In certain embodiments, the permeable support is at least a gas-permeable membrane.

[0009] In certain embodiments, the permeable support is a porous surface that allows gas transfer between the first and second reservoirs.

[0010] In certain embodiments, the permeable support is a porous surface that allows passage of at least one or more of gases, nutrients, and metabolites between the first and second reservoirs.

[0011] In certain embodiments, the seal prevents mixing of the first and second gaseous environments except through the permeable support. In this manner, the device provides two separate gaseous atmospheres within the first and second chambers, which are connected only through the permeable support for purposes of gas and / or nutrient transfer between the two chambers. In this manner, the biological modeling device includes a sealing mechanism for maintaining sealed, separate (i.e., separately controllable) gaseous environments within the first and second reservoirs. This configuration allows for the introduction and sampling of multiple chambers with separate gaseous atmospheres without disturbing or intermixing the gaseous atmospheres. Gas can move through the permeable support. There is diffusion / migration of gas through biological elements supported by the permeable support, such as through biological materials / cells. By providing two gaseous environments sealed from each other except through the permeable support, the device is operable to biologically model conditions having anaerobic and aerobic separation and / or test smoke, airborne toxins, drugs, etc. across biological elements such as colon cell populations, epithelial cell populations, in vitro airway models, etc., held on the permeable support.

[0012] As referred to herein, a "main path" or "primary path" is a path through which substantially all of the gas movement occurs. In certain embodiments, the gas flow between the first reservoir and the second reservoir passes through the permeable support as the main path. Any inherent gas permeability of the seal material is small compared to the gas permeability of the permeable support.

[0013] In certain embodiments, the first gaseous environment comprises a headspace of a first reservoir of the first chamber.

[0014] In certain embodiments, the second gaseous environment comprises a headspace of a second reservoir of the second chamber.

[0015] As referred to herein, the "headspace" of a reservoir is the gas space above the biological component and / or any solution within the reservoir.

[0016] In certain embodiments, the biological modeling device is useful for co-culturing cells. In such a system, each of the first and second chambers contains a biological element in the form of one or more cell populations, and the main pathway between the chambers is through the permeable support. In particular, a flow path is provided through the permeable support as the main (i.e., primary) pathway. One particular advantage of this configuration is that, for example, an aerobic environment can be used to grow one cell population, while an anaerobic environment can simultaneously be used to grow another cell population. Thus, each environment can be individually controlled independently of the other environment.

[0017] In certain embodiments, the second gaseous environment is sealed from the first gaseous environment by a combination of a seal and the first chamber.

[0018] In certain embodiments, the first gaseous environment is an anaerobic environment.

[0019] In certain embodiments, the second gaseous environment is an aerobic environment.

[0020] In certain embodiments, the first and / or second reservoirs are configured to receive and retain at least one biological component, which may be one or more of a mixed cell population, a single cell population, a co-culture of cells, a bacterial culture, a microbial population, mucus, etc.

[0021] Additionally or alternatively, the first and / or second reservoirs are configured to receive and hold a solution, which in certain embodiments may be one or more of a culture or nutrient medium, a buffer, a test solution, etc.

[0022] In certain embodiments, the seal is a gasket seal.

[0023] In certain embodiments, the permeable support is integrally formed with the gasket seal. In such embodiments, a separate first chamber element is not required, and the gasket seal forms the first chamber.

[0024] In certain embodiments, the gasket seal comprises a pierceable portion.

[0025] In certain embodiments, the puncturable portion comprises a septum.

[0026] In certain embodiments, the septum is a resealable septum or a self-sealing septum. In this way, a sampling or injection device can be introduced into the first or second chamber independently of the other chamber. The gas environment of the chamber receiving the sampling or injection device remains sealed and separated from the gas environment of the other chamber.

[0027] In certain embodiments, the first chamber is an insert comprising a permeable support.

[0028] In certain embodiments, the permeable support forms at least a portion of the bottom surface of the first chamber.

[0029] In certain embodiments, the insert comprises a wall upstanding from the bottom surface of the permeable support.

[0030] In certain embodiments, the first chamber is partially received into the second chamber through a gasket seal.

[0031] In certain embodiments, the seal comprises a barrier layer disposed in sealable engagement with the second chamber to separate the second gaseous environment from the first gaseous environment.

[0032] In certain embodiments, the barrier layer comprises an opening.

[0033] In certain embodiments, the first chamber is partially received through the opening such that a sealing engagement is formed between the barrier layer and the first chamber, thereby sealing the second chamber with the combination of the barrier layer and the first chamber.

[0034] In certain embodiments, the opening in the barrier layer is a self-sealing septum such that when a first chamber is forced through the opening, the barrier layer forms a sealing engagement with the first chamber.

[0035] In certain embodiments, the first chamber is part of the barrier layer.

[0036] In certain embodiments, the first chamber is integrally formed with the barrier layer.

[0037] In certain embodiments, the permeable support is integrally formed with the barrier layer. In such embodiments, a separate first chamber element is not required, and the barrier layer forms the first chamber.

[0038] In certain embodiments, the second chamber is located within the base plate. In certain embodiments, the second chamber is integrally formed with the base plate.

[0039] In certain embodiments, the second chamber comprises a bottom surface and an inner wall extending upward from the bottom surface to an open top, the bottom surface and inner wall cooperatively defining a second reservoir.

[0040] In certain embodiments, the biological modeling apparatus includes a plurality of second chambers, each of the second chambers including a second reservoir.

[0041] In certain embodiments, the biological modeling device comprises a plurality of first chambers, each of which corresponds to a respective one of a plurality of second chambers, and each of which comprises a first reservoir and a permeable support positioned between the respective first and second reservoirs.

[0042] In certain embodiments, each first chamber is an insert.

[0043] In certain embodiments, each first chamber is integrally formed with a seal and each first chamber is integrally formed with a permeable support.

[0044] In certain embodiments, the biological modeling apparatus includes a plurality of seals, each seal disposed between and sealably engaging a respective first chamber and second chamber.

[0045] In certain embodiments, the seal is disposed between an outer surface of the first chamber and an inner wall of the second chamber and sealably engages the outer surface of the first chamber and the inner wall of the second chamber.

[0046] In certain embodiments, the biological modeling apparatus further comprises a cover for the or each first chamber, the first chambers being closed by the cover.

[0047] In certain embodiments, the cover comprises a seal disposed in sealable engagement with the first chamber to separate the first gaseous environment from the external ambient environment, the first chamber being sealed by the seal.

[0048] In certain embodiments, the seal of the cover comprises a pierceable portion.

[0049] In certain embodiments, the puncturable portion comprises a septum.

[0050] In certain embodiments, the septum is a resealable septum or a self-sealing septum.

[0051] In certain embodiments, the cover seal includes two or more pierceable portions, preferably two pierceable portions.

[0052] In certain embodiments, one puncturable portion is configured to allow access to one of the first chamber or the second chamber, and another of the puncturable portions is configured to allow access to the other of the first chamber and the second chamber.

[0053] In certain embodiments, each pierceable portion comprises a septum.

[0054] In certain embodiments, the septum is a resealable septum or a self-sealing septum.

[0055] In certain embodiments, the biological modeling apparatus further comprises a transfer cover. More specifically, the transfer cover is formed of a gas permeable material. Even more specifically, the transfer cover is constructed and arranged to cover at least one of the first chamber(s), or each first chamber. In this manner, the biological component(s) and / or solution(s) in the first chamber(s) are prevented from transferring between adjacent first chambers.

[0056] In certain embodiments, the transmission cover is a gas permeable silicone membrane.

[0057] In certain embodiments, the first chamber is the apical chamber.

[0058] In certain embodiments, the second chamber is a basolateral chamber.

[0059] In certain embodiments, the first reservoir is configured to receive and hold a first culture medium for delivery to the first biological component, and in certain embodiments, the second reservoir is configured to receive and hold a second culture medium for delivery to the first and / or second biological component.

[0060] In certain embodiments, the second reservoir is configured to receive and retain a second culture medium for maintaining the first biological component through the permeable support. More specifically, the second culture medium can provide nutrients to the first biological component through the permeable support.

[0061] In certain embodiments, the first chamber comprises a sealing plug operable to seal the first reservoir from the ambient environment, in this manner the first chamber may be closed and unused.

[0062] In some embodiments, the biological modeling apparatus is a culture apparatus for growing cells and / or tissues in vitro.

[0063] According to another aspect of the present disclosure, there is provided a method of growing cells and / or tissue in vitro using a biological modeling apparatus according to another aspect of the present invention.

[0064] According to a further aspect of the present disclosure, there is provided a biological modeling apparatus, the apparatus comprising: a first chamber providing a first reservoir having a first gas environment and a second chamber having a second gas environment, the second reservoir and the first reservoir being connected via a permeable support, and the first chamber being at least partially received within the second chamber; This device is a seal disposed to separate a first gaseous environment of the first reservoir in the first chamber from a second gaseous environment of the second reservoir in the second chamber and sealably engages the first and second chambers; a cover for the first chamber; Equipped with The first chamber is closed by a cover, the cover having at least one opening and a duct extending from the at least one opening towards the first chamber.

[0065] In certain embodiments, the cover further comprises at least one opening and a duct extending from the at least one opening toward the second chamber.

[0066] In certain embodiments, the first reservoir is configured to receive and hold a first biological component.

[0067] In certain embodiments, the second reservoir is configured to receive and hold a second biological component.

[0068] In certain embodiments, the first and / or second biological component may comprise a cell population. More specifically, the cell population may be selected from one or more of bacteria, epithelial cells, and immune cells.

[0069] In certain embodiments, the first and / or second reservoirs receive and hold a solution. More specifically, the solution may be one or more of a culture or nutrient medium, a buffer, a test solution, and the like.

[0070] According to another aspect of the present disclosure, there is provided a biological modeling apparatus, the apparatus comprising: a first chamber providing a first reservoir having a first gaseous environment and a second chamber providing a second reservoir having a second gaseous environment, the second reservoir and the first reservoir being connected via a permeable support, and the first chamber being at least partially received within the second chamber; This device is a seal disposed to separate a first gaseous environment of the first reservoir in the first chamber from a second gaseous environment of the second reservoir in the second chamber and sealably engages the first and second chambers; a cover for the first chamber; Equipped with The first chamber is closed by a cover, the cover having at least one opening and a duct extending from the at least one opening towards the second chamber.

[0071] In certain embodiments, the cover further comprises at least one opening and a duct extending from the at least one opening toward the first chamber.

[0072] In certain embodiments, the first reservoir is configured to receive and hold a first biological component.

[0073] In certain embodiments, the second reservoir is configured to receive and hold a second biological component.

[0074] In certain embodiments, the first and / or second biological component may comprise a cell population. More specifically, the cell population may be selected from one or more of bacteria, epithelial cells, and immune cells.

[0075] In certain embodiments, the first and / or second reservoirs receive and hold a solution. More specifically, the solution may be one or more of a culture or nutrient medium, a buffer, a test solution, and the like.

[0076] Embodiments of the present invention will now be described, by way of example only, with reference to the accompanying drawings, in which: [Brief explanation of the drawings]

[0077] [Figure 1a] 1 shows an example of a biological modeling device in an exploded perspective view. [Figure 1b] 1 shows an example of a biological modeling device in an exploded front perspective view. [Figure 1c] 1 shows an assembled perspective view of an example biological modeling device. [Figure 1d]1 shows an example of a biological modeling device in a cross-sectional front view. [Figure 1e] 1 shows an example of a biological modeling device in an enlarged cross-sectional view from the front. [Figure 2a] 1 illustrates another example of a biological modeling device in an exploded perspective view. [Figure 2b] 1 illustrates another example of a biological modeling device in an exploded front perspective view. [Figure 2c] 1 illustrates another example of a biological modeling device in an assembled perspective view. [Figure 2d] 1 shows another example of a biological modeling device in a front cross-sectional view. [Figure 2e] 1 shows another example of a biological modeling device in an enlarged cross-sectional front view. [Figure 3a] 3 illustrates an exploded perspective view of the biological modeling device of FIG. 2 further including a sheet and a puncturable film substrate. [Figure 3b] 3 illustrates an exploded front perspective view of the biological modeling device of FIG. 2 further including a sheet and a puncturable film substrate. [Figure 3c] 3 illustrates the assembled perspective view of the biological modeling device of FIG. 2, further including a sheet and a puncturable film substrate. [Figure 3d] 3 shows the biological modeling device of FIG. 2 in a front cross-sectional view, further including a sheet and a puncturable film substrate. [Figure 3e] 3 shows an enlarged cross-sectional front view of the biological modeling device of FIG. 2 further including a sheet and a puncturable film substrate. [Figure 4a] 1 illustrates another example of a biological modeling device in an exploded perspective view from above. [Figure 4b] 1 shows another example of a biological modeling device in an exploded perspective view from below. [Figure 4c] 1 shows another example of a biological modeling device in an assembled cross-sectional view. [Figure 5a]10 shows a further example of a biological modeling device from a close-up of the insert in an exploded perspective view. [Figure 5b] 10 shows a further example of a biological modeling device from a close-up of the insert in a partially exploded perspective view. [Figure 5c] 10A and 10B show further examples of biological modeling devices from a close-up of the insert in an assembled perspective view. [Figure 5d] 10A and 10B show further examples of biological modeling devices from a close-up of the insert in an assembled cross-section. [Figure 6a] 1 shows a perspective view of a portion of a base plate. [Figure 6b] 1 shows a cross-sectional view of a portion of a base plate. [Figure 7a] 1 shows the baseplate of a biological modeling device printed using fused deposition modeling (FDM). [Figure 7b] 1 shows the baseplate of a biological modeling device printed using stereolithography (SLA). [Figure 8a] 1 shows a perspective view of the base plate of the biological modeling device. [Figure 8b] 1 shows a perspective view of the insert tray of the biological modeling device. [Figure 8c] 1 shows a perspective view of an assembly of a base plate and an insert tray. [Figure 8d] 1 shows a perspective view of an assembly of a base plate, an insert tray, and a cover. [Figure 8e] 1 shows a cross-sectional view of an assembly including a base plate and an insert tray. [Figure 9a] 1 shows a model of a biological modeling device fabricated using stereolithography 3D printing with a separate base plate, insert tray, and cover. [Figure 9b]1 shows a model of a biological modeling device fabricated using stereolithography 3D printing with an assembled base plate and insert tray and a separate cover. [Figure 9c] 1 shows a model of a biological modeling device fabricated using stereolithography 3D printing with an assembly of a base plate, an insert tray, and a cover. [Figure 10] 1 illustrates a biological modeling apparatus comprising a transfer cover constructed and arranged to prevent mass transfer between adjacent first chambers. DETAILED DESCRIPTION OF THE INVENTION

[0078] Certain terminology is used in the following description for convenience only and not as a limitation. The words "right," "left," "lower," "upper," "front," "rear," "upward," "down," and "downward" designate directions in the drawing figures to which reference is made and relate to the described components in their assembled and mounted state. The words "inner," "inwardly," and "outer," "outwardly" refer to directions toward and away from a designated centerline or geometric center (e.g., central axis) of the described element, respectively, with the particular meaning being readily apparent from the context of the description.

[0079] Furthermore, as used herein, the terms "connected," "attached," "coupled," and "mounted" are intended to include a direct connection between two elements with no other intervening elements present, and an indirect connection between elements with one or more intervening elements present. The terminology includes the words specifically mentioned above, derivatives thereof, and words of similar import.

[0080] Furthermore, unless otherwise specified, the use of ordinal adjectives such as "first," "second," "third," etc., merely indicates reference to different instances of similar objects and does not imply that the objects so described must be in a given order in time, space, ranking, or in any other way.

[0081] Like reference numbers are used throughout to refer to like features.

[0082] Referring now to FIG. 1, a biological modeling apparatus 2 is shown. The biological modeling apparatus 2 includes several inserts (i.e., first chambers) 4 that are inserted through openings in a tray 27 in this example. In the illustrated embodiment, the inserts 4 include outwardly extending arms 5 that engage the surface of the tray 27 to hold the inserts 4 in place. Each insert 4 has a tubular inner wall 8 that terminates in a bottom surface 6 located at one end and an open top located at the other end. The tray 27 forms an airtight seal around each insert 4. A porous membrane 20 is integrally formed on the bottom surface 6 of each insert 4. The inserts 4 define reservoirs for holding biological material (e.g., cell populations) to be cultured, such as a population containing bacterial cells. In this example, the inserts 4 are apical chambers. The biological modeling apparatus 2 is suitable for in vitro cell and / or tissue culture. The pores in the membrane 20 are between 0.4 and 8 microns in size. In one particular example, a membrane 20 with a pore size of 1.8 microns is used to model the intestine. The membrane 20 has a scaffold structure that promotes tissue-like behavior of the cultured cells. The scaffold structure may be, for example, a gel or other tissue culture matrix.

[0083] The device 2 includes wells (i.e., second chambers) 12, each formed as part of a base plate 24. The number of wells 12 corresponds to the number of inserts 4 in the tray 27. In this example, 24 wells 12 are provided, arranged in a 6x4 matrix; however, any number of wells 12 is contemplated depending on cell growth requirements, such as 6, 12, 24, 48, 96, 384, or 1536 wells 12. Each well 12 has a tubular inner wall 16 terminating in a bottom 14 at one end and an open top at the other end. The inserts 4 are positioned within the tray 27 in alignment with corresponding wells 12 in the base plate 24, such that the central axis of each insert 4 is aligned with the central axis of each well 12. The inserts 4 have a diameter smaller than the diameter of the corresponding tubular well 12. Each of the wells 12 defines a reservoir for holding biological material, such as a population containing epithelial cells. The insert 4 and well 12 in this example are tubular in shape, although other regular and irregular shapes are envisioned. In this example, the well 12 is a basolateral chamber.

[0084] A seal 22 is provided surrounding the or each insert 4. In this example, an individual seal 22 is provided surrounding each insert 4. The seals 22 are sized so that the inserts 4 of the tray 27 are received through the seal 22 and placed within the wells 12 of the base plate 24. When the inserts 4 are received in their respective wells 12 in this manner, the seals 22 engage the inserts 4 and the wells 12 to provide a sealing mechanism that separates the gaseous environment within the headspace of the insert 4 from the gaseous environment within the headspace of the wells 12. Specifically, the seals 22 are disposed between the outer surface of the inserts 4 and the interior walls 16 of the wells 12. The headspace 18 of the wells 12 is closed by the seals 22 and the inserts 4, thereby providing a seal. In this particular example, a gasket 23 is provided to improve the seal between the inserts 4 and the wells 12, as best seen in FIG. 1(e).

[0085] The base plate 24 includes a skirt 26 that extends continuously around the outer edge of the base plate 24. The tray 27 includes a flange 28 that extends outward from the tray 27. The tray 27 is sized and shaped so that the flange 28 of the tray 27 is retained within the base plate 24 by the skirt 26 of the base plate 24. With the tray 27 retained within the base plate 24 in this manner, each insert 4 is aligned with and inserted toward a respective well 12. At the same time, the seal 22 contacts the inner wall 16 of the well 12, providing a sealed environment within the reservoir of the well 12. The seal 22 is formed with a barrier layer 30 that cooperates with the insert 4 to seal the well 12. One surface of the base plate 24 is provided with an air valve 25 that allows the free flow of air from the ambient environment outside the device 2 into the base plate 24, and more specifically, into the wells 12. This maintains an aerobic environment within the wells 12. It is contemplated that the air valve 25 may be provided with a particulate filter.

[0086] The or each insert 4's reservoir is connected to the respective well 12 via a porous membrane 20. While the porous membrane 20 is integrally formed with the bottom surface 6 in this example, it is contemplated that the porous membrane 20 may be provided as a separate component disposed on the bottom surface 6 of the insert 4. The porous membrane 20 may be provided in the form of a mesh or matrix of holes. The porosity of the membrane 20 allows, for example, the passage of gases, nutrients, and metabolic products between the insert 4 and the respective reservoirs of the well 12.

[0087] The biological modeling apparatus 2 includes a cover 32 that encloses the headspace 10 of the insert 4 when in the closed position. The lower portion of the cover 32 includes a flange 34 that extends around the outer edge of the cover 32. A seal 38 is positioned to engage the flange so that the cover 32 is sealed against the tray 27 and base plate 24 when the cover 32 is closed. The seal 38 may be a gasket seal. Each of the outwardly facing surfaces of the cover 32 has a latch 36 attached to the surface of the cover 32 via a hinge, and each latch 36 can pivot independently relative to the cover 32 about its respective hinge.

[0088] 1(c)-1(e), in use, each insert 4 is supplied with a first biological component (e.g., bacterial cells) and a maintenance medium, such as phosphate-buffered saline (PBS). Each well 12 is supplied with a second biological component (e.g., epithelial cells) and a different medium for culturing the cells. The tray 27 is lowered onto the base plate 24 so that each insert 4 is lowered into its respective well 12, with the porous membrane 20 interposed between the insert 4 and the reservoir of the well 12. When the tray 27 is fully received within the base plate 24, the flange 28 of the tray 27 is retained by the skirt 26 of the base plate 24. In this configuration, the seal 22 engages with the inner wall 16 of the well 12 and the insert 4, thereby closing the headspace 18 of the well 12. In this manner, the gaseous environment within the headspace 10 of the insert 4 and the gaseous environment within the headspace 18 of the well 12 are separated by the seal 22. At the same time, the porous membrane 20 provides a pathway for the transfer of gases, nutrients, and / or metabolites between the reservoirs of the respective inserts 4 and wells 12.

[0089] When the tray 27 and base plate 24 are assembled together to receive the inserts 4 within the wells 12, the headspace 10 of each insert 4 is open to the ambient environment. To close the headspace 10 of the inserts 4 from the environment, a cover 32 is placed over the tray 27 and latches 36 are pivoted about their respective hinges to engage the bottom surface of the skirt 26.

[0090] The lower chamber (i.e., well 12) can be referred to as the basolateral chamber, and the upper chamber (i.e., insert 4) can be referred to as the apical chamber. In use, the upper chamber can contain bacterial cells cultured in an anaerobic environment. A sachet can be placed within the chamber to create the anaerobic environment. The contents of the sachet initiate a reaction that suppresses oxygen in the environment. The lower chamber can contain mammalian epithelial cells cultured in an aerobic environment. In this example, the aerobic environment can be provided by an air valve 25, which allows free flow of air into the lower chamber. However, in other examples, both the upper and lower chambers could be supplied with gas from an external source. Thus, the gas environment of the upper chamber is separated from that of the lower chamber, providing a controlled way to simultaneously culture cells in separate gas environments. The upper chamber can be used to model the lumen. The lower chamber can be used to model the intestinal wall. It is contemplated that a mucus layer may additionally be formed on the porous membrane 20. In particular, it is contemplated that a mucus layer may be formed on epithelial cells to simulate the intestinal epithelium. It is also contemplated that when some inserts 4 are not used, the open tops of the inserts 4 may be sealed with sealing plugs (not shown). In this way, the redundant inserts 4 are closed and not used.

[0091] FIG. 2 shows another example of a biological modeling apparatus 2 in which wells 12, seals 22, and covers 32 differ from the wells, seals, and covers of the example shown in FIG. 1 . The biological modeling apparatus 2 includes a plurality of wells 12 formed as part of a base plate 24. Each well 12 in this embodiment has an interior wall 16 having a square or rectangular footprint, terminating in a bottom surface 14 and an open top opposite the bottom surface 14. Each well 12 defines a reservoir for holding biological material, such as epithelial cells. An insert 4 is provided in the biological modeling apparatus 2 in the same manner as in FIG. 1 , and therefore will not be described in detail again here.

[0092] A seal 22 is provided surrounding the or each insert 4 such that the insert 4 is received through the seal 22 and positioned within the well 12 of the base plate 24. The seal 22 in this example has a frustoconical profile that tapers inward toward the base plate 24. In use, when the tray 27 is received on the base plate 24, the insert 4 lowers into the respective well 12. As the insert 4 is lowered into the well 12 in this manner, the lower end of the seal 22 engages the inner wall 16 of the well 12, and the headspace 18 of the well 12 is closed and sealed by the seal 22 and the respective insert 4. It is contemplated that the seal 22 may have a pierceable portion. The pierceable portion may have a resealable septum. In certain examples, the pierceable portion of the seal 22 may be a self-sealing septum.

[0093] The biological modeling device 2 includes a cover 32 that encloses the head space 10 of the insert 4 when in the closed position. The cover 32 in this example differs from the embodiment of FIG. 1 in that it includes an array of first openings 40 and second openings 42. The diameter of the first openings 40 is larger than the diameter of the second openings 42. The first openings 40 are arranged in a 6x4 matrix aligned with the central axis of each insert 4. Each of the second openings 42 is offset from the first opening 40 and, therefore, from the central axis of each insert 4. Each of the first openings 40 is covered by a puncturable film 41. Similarly, each of the second openings 42 is covered by a puncturable film 43. The puncturable films 41, 43 have resealable partitions. More specifically, the puncturable films 41, 43 have self-sealing partitions.

[0094] As shown in FIGS. 2(d) and 2(e), each of the first openings 40 leads to a duct 44 that provides a passageway toward the respective aligned insert 4 for introducing / extracting medium and / or cells into / from the insert 4 when the film 41 is punctured. In this example, each of the second openings 42 leads to another duct 46 that provides a passageway toward the respective well 12 for introducing / extracting medium and / or cells into / from the well 12 when the film 43 is punctured. In this example, the ducts 44, 46 are shown extending linearly downward, but in other examples, the ducts 44, 46 may instead extend in different directions toward the insert 4 and / or well 12 or may extend non-linearly. For example, the ducts 44, 46 may extend at an angle offset by 20 degrees from the central axis of the well 12. In a configuration not shown, the ducts 44, 46 extend into the insert 4 and function as condensation collectors. In this way, condensation can be collected while preventing the transfer of biological materials, such as bacteria, between adjacent inserts 4. Therefore, the sterility of the device can be improved. The gas permeability of the ducts 44, 46 ensures a continuous first gas environment between adjacent upper chambers (e.g., inserts 4).

[0095] While in this example, well 12, seal 22, and cover 32 have each been modified from the example of Figure 1, other examples are contemplated in which one or more, but not all, of well 12, seal 22, and cover 32 are modified from the example of Figure 1. For example, it is contemplated that biological modeling apparatus 2 may include all of the features of Figure 1, with cover 32 modified to include openings 40, 42 and respective puncturable films 41, 43. Another example of biological modeling apparatus 2 may include cover 32 with openings 40, 42, puncturable films 41, 43, and ducts 44, 46.

[0096] FIG. 3 illustrates a further example of a biological modeling device 2 that is substantially similar to device 2 of FIG. 2. However, larger first opening 40 is not sealed by puncturable film 41. Similarly, smaller second opening 42 is not sealed by puncturable film 43. Rather, openings 40, 42 are through-holes extending through the entire thickness of cover 32. The biological modeling device 2 of FIG. 3 includes a sheet 50 having openings 52, 54 positioned to align with larger first opening 40 and smaller second opening 42, respectively, of cover 32. Specifically, openings 52 in sheet 50 are appropriately sized and spaced to align with larger first opening 40 in cover 32. Openings 54 in sheet 50 are appropriately sized and spaced to align with smaller second openings 42 in cover 32. A puncturable substrate layer 48 is disposed between cover 32 and sheet 50. The puncturable substrate layer 48 is sized and positioned to cover and extend between the openings 40 , 42 in the cover 32 and the openings 52 , 54 in the sheet 50 .

[0097] In this example, the puncturable substrate layer 48 has a resealable septum. It is contemplated that the puncturable substrate layer 48 may have a self-sealing septum. In use, the substrate layer 48 is punctured through the opening 52 to access the lower insert 4 through the duct 44 for introducing / extracting medium and / or cells into / from the insert 4. The substrate layer 48 is punctured through the opening 54 to access the lower well 12 through the duct 46 for introducing / extracting medium and / or cells into / from the well 12.

[0098] FIG. 4 shows an embodiment of a biological modeling apparatus 2 having an alternative sealing configuration to the previous example. Each insert 4 in a tray 27 is provided with a seal in the form of an O-ring 22 that is received around the insert 4 and abuts the underside of the tray 27. When the insert 4 is received in a corresponding well 12 in a base plate 24, the chamber of the well 12 is separated from the chamber of the insert 4. The O-ring 22 provides a fluid-tight and gas-tight seal between the insert 4 and the well 12. The base plate 25 includes an air valve 25 that may be connected to an external tube or pipe to introduce gas or fluid into the well 12. The O-ring 22 is located above the chamber of the well 12, allowing gas to pass through the entire base plate 24 and into the interior of the well 12 during use. In this example, the O-ring 22 is made of nitrile rubber and has an inner diameter of 14 mm and an outer diameter of 18 mm.

[0099] FIG. 5 shows a further variation of the biological modeling apparatus 2 in which the porous membrane 20 is attached to the bottom of the insert 4 and held in place by clips 58. The insert 4 includes protrusions 56 on either side of its underside. The protrusions 56 engage opposing passages (i.e., notches) 60 in the clip, holding the clip in place around the outer wall of the insert 4. The porous membrane 20 is sized to be larger than the opening of the clip 58 so that the porous membrane 20 is pressed against the insert 4 and held in place. By providing the clip 58 in this manner, no adhesive is required to attach the porous membrane 20 in place. In this example, the gap between the bottom of the clip 58 and the bottom of the well 12 is 2 millimeters (mm).

[0100] 6 illustrates an alternative embodiment showing a portion of a base plate 24 having a single well 12. The well 12 includes a recess (i.e., slot) 62 at its distal end that receives an O-ring seal. By providing an O-ring within the recess 62, a seal is formed between the base plate 24 and an insert received within the well 12.

[0101] Figure 7 shows two different samples of base plate 24 manufactured using different 3D printing techniques. Figure 7(a) shows a base plate 24 3D printed using fused deposition modeling (FDM) with acrylonitrile butadiene styrene (ABS) as the printing material. Figure 7(b) shows a base plate 24 3D printed using stereolithography (SLA) with Biomed clear resin as the printing material.

[0102] Figure 8 shows the various components of the biological modeling apparatus. Figure 8(a) shows a base plate 24 having eight tubular wells 12 and valves 25 at either end for introducing gas or fluid into the wells 12 during cell culture. Figure 8(b) shows a tray 27 containing eight inserts 8 shaped and sized to be held within the wells 12. The base plate 24 and tray 27 are assembled together during use, and may be provided with a cover 32 to prevent evaporation of contents within the apparatus, as shown in Figure 8(d).

[0103] As shown in FIG. 9 , each of the base plate 24, insert tray 27, and cover 32 may be separately manufactured (e.g., by 3D printing). In this particular example, the components are printed using stereolithography 3D printing, using acrylonitrile butadiene styrene (ABS) as the printing material. To assemble the biological modeling apparatus 2, the tray 27 is placed on the base plate 24 so that the inserts 4 are held within the corresponding wells 12. A cover 32 is then placed over the assembled tray 27 and base plate 24 to prevent evaporation of the internal contents during cell culture. The base plate 24 includes a valve 25 that may be attached to an external pipe or tube (not shown) to introduce fluid or gas into the chambers of the wells 12. For example, oxygen can be introduced through the valve 25 to create an aerobic environment within the wells 12.

[0104] 10 illustrates a biological modeling apparatus 2 having a gas-permeable silicone transfer cover 64. Transfer cover 64 sits on top of insert tray 27 and prevents transfer of biological material and / or solutions between adjacent inserts 4. The gas permeability of transfer cover 64 ensures a continuous first gas environment between adjacent upper chambers (e.g., inserts 4).

[0105] The biological modeling apparatus of the present invention is suitable for use in several exemplary models. The following models and biological materials are presented as examples:

[0106] Human colon model A representative colon cell line (such as CACO-2 or T84) is grown in monolayer on a transwell insert 4. A cell-compatible mucus layer is placed on top of the cell layer in the first chamber provided by the insert 4. A bacterial inoculum and nutrient medium are placed on top of the mucus layer. The bacterial inoculum can be a single bacterium, multiple bacteria, or one of a representative microbial culture grown from a fecal inoculum.

[0107] The insert 4 provides an upper chamber with an anaerobic environment (less than about 0.5-1% oxygen) generated by the GasPak™. The lower chamber includes a well 12 in which an aerobic environment is provided. The lower chamber can hold a nutrient medium and may hold a mixed cell population including immune cells, fibroblasts, epithelial cells, and / or blood cells.

[0108] Samples can be taken from either chamber (insert 4 and / or well 12) throughout the modeling regime, or at the end or at specified time points. Samples can be analyzed for metabolites, mass transfer, 16s sequencing of bacteria, RNA sequencing from cells, etc.

[0109] Cell-free colon model This model is similar to the human colon model described above without the inclusion of representative colon cell lines (such as CACO-2 or T84).

[0110] Bacteria-free colon model This model is similar to the human colon model described above, in that the biological modeling apparatus is kept in a sterile environment and is not inoculated with bacteria.

[0111] Airway Mode Representative airway cell lines (such as Calu-3 or primary airway cultures) are grown in monolayer on the permeable support of the transwell insert 4. A cell-compatible mucus layer is placed on top of the cell layer to represent airway mucin in the model. Bacteria may be introduced in certain instances of the model. The headspace of the apical chamber is an air space containing gas. Nutrient medium is provided to the basolateral chamber.

[0112] In this model, both chambers are filled with an aerobic gas environment to simulate lung function. The gas environment within the two chambers can be adjusted to mimic conditions such as hypercapnia, hyperoxemia, cigarette smoke, pollution, and carbon monoxide poisoning.

[0113] Mucus-free airway model This model is similar to the airway model described above: in this model, the mucus layer is not present.

[0114] Those skilled in the art will appreciate that the above detailed examples are set forth by way of example only and not in a limiting sense, and that various changes and modifications may be made without departing from the scope of the invention as defined by the appended claims. Various modifications to the above detailed examples are possible.

[0115] Throughout the description and claims of this specification, the terms "comprise" and "contain" and variations thereof mean "including but not limited to" and are not intended to (and do not) exclude other moieties, additives, components, integers, or steps. Throughout the description and claims of this specification, the singular includes the plural unless the context otherwise requires. In particular, where the indefinite article is used, the specification should be understood as contemplating the plural as well as the singular unless the context otherwise requires.

[0116] It should be understood that any feature, integer, property, compound, chemical moiety, or group described in connection with a particular aspect, embodiment, or example of the invention is applicable to any other aspect, embodiment, or example described herein, unless incompatible. All features disclosed herein (including the accompanying claims, abstract, and drawings) and / or all steps of any method or process so disclosed may be combined in any combination, except combinations in which at least some of such features and / or steps are mutually exclusive. The invention is not limited to the details of any of the embodiments described above. The invention extends to any novel one or any novel combination of features disclosed herein (including the accompanying claims, abstract, and drawings), or any novel one or any novel combination of steps of any method or process so disclosed.

[0117] Those skilled in the art will appreciate that the above embodiment(s) have been described by way of example only, and not by way of limitation, and that various changes and modifications may be made without departing from the scope of the invention as defined by the appended claims. Various modifications may be made to the detailed designs set forth above. [Explanation of symbols]

[0118] Table 1

Claims

1. a first chamber providing a first reservoir having a first gaseous environment and a second chamber providing a second reservoir having a second gaseous environment, the first and second reservoirs being connected via a permeable support, and the first chamber being at least partially received within the second chamber; a seal disposed to separate a first gaseous environment of the first reservoir from a second gaseous environment of the second reservoir and sealably engage the first and second chambers; The biological modeling device further comprises:

2. The biological modeling apparatus of claim 1 , wherein the permeable support is a membrane, which may be a gas-permeable membrane.

3. 10. The biological modeling apparatus of claim 1, wherein the first chamber is configured to accommodate and hold a first biological element and / or solution, and the second chamber is configured to accommodate and hold a second biological element and / or solution.

4. 4. The biological modeling apparatus of claim 1, wherein the first gas environment comprises a headspace of the first reservoir in the first chamber, and the second gas environment comprises a headspace of the second reservoir in the second chamber.

5. The biological modeling apparatus of claim 1 , wherein the second gaseous environment is sealed from the first gaseous environment by a combination of the seal and the first chamber.

6. The biological modeling apparatus of any one of claims 1 to 5, wherein the seal is a gasket seal.

7. The biological modeling apparatus of claim 6 , wherein the gasket seal comprises a puncturable septum, which may be a self-sealing septum.

8. 8. The biological modeling apparatus of claim 6 or 7, wherein the first chamber is partially received into the second chamber through the gasket seal.

9. 9. The biological modeling apparatus of claim 1, wherein the seal comprises a barrier layer disposed in sealable engagement with the second chamber to separate the second gaseous environment from the first gaseous environment, the barrier layer comprising an opening, the first chamber being partially received through the opening such that a sealing engagement is formed between the barrier layer and the first chamber, thereby sealing the second chamber by the barrier layer and the first chamber.

10. The biological modeling apparatus of any one of claims 1 to 9, wherein the second chamber is located within a base plate or is formed as part of a base plate.

11. 11. The biological modeling device of claim 1, wherein the second chamber comprises a bottom surface and an inner wall extending upward from the bottom surface toward an open top, the bottom surface and inner wall cooperatively defining the second reservoir.

12. The biological modeling apparatus of any one of claims 1 to 11, wherein the first chamber is an insert comprising the permeable support.

13. The biological modeling apparatus of claim 12 , wherein the permeable support forms at least a portion of the bottom surface of the first chamber.

14. The biological modeling apparatus of any one of claims 1 to 13, comprising a plurality of second chambers, each second chamber comprising a second reservoir.

15. 15. The biological modeling apparatus of claim 14, comprising a plurality of first chambers, each first chamber being an insert corresponding to each of the plurality of second chambers, each insert comprising a first reservoir and a permeable support positioned between the respective first and second reservoirs.

16. 16. The biological modeling apparatus of claim 15, comprising a plurality of seals, each seal disposed between and in sealable engagement with the respective first and second chambers.

17. The biological modeling apparatus of any one of claims 1 to 16, wherein the first chamber may be an apical chamber and the second chamber may be a basolateral chamber.

18. 18. The biological modeling apparatus of claim 1, wherein the seal is disposed between an outer surface of the first chamber and an inner wall of the second chamber and sealably engages the outer surface of the first chamber and the inner wall of the second chamber.

19. 19. The biological modeling apparatus of claim 1, further comprising a cover for the first chamber, the head space of the first chamber being closed by the cover.

20. 20. The biological modeling apparatus of claim 19, wherein the cover comprises a seal disposed in sealable engagement with the first chamber to separate the first gaseous environment from an external ambient environment, the headspace of the first chamber being sealed by the seal.

21. The biological modeling apparatus of any one of claims 1 to 20, wherein the seal of the cover comprises at least one pierceable portion.

22. 22. The biological modeling apparatus of claim 21 , wherein the seal of the cover comprises at least two puncturable portions, one puncturable portion configured to allow access to one of the first chamber or the second chamber, and another of the puncturable portions configured to allow access to the other of the first chamber and the second chamber.

23. 23. The biological modeling apparatus of claim 21 or 22, wherein the puncturable portion comprises a septum, which may be a resealable septum or a self-sealing septum.

24. 24. The biological modeling device of any one of claims 1 to 23, which is a culture device for growing cells and / or tissues in vitro.

25. A method for the co-cultivation of cells using a biological modelling device according to any one of claims 1 to 24.

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