Device for simulating a biological barrier

Abstract

A device for simulating a biological barrier comprising a multi-well plate covered by a port plate and sustained by a connection plate ensuring connection to a biochip.

Description

DEVICE FOR SIMULATING A BIOLOGICAL BARRIERFIELD OF INVENTION

[0001] The present invention relates to a device for simulating a biological barrier.BACKGROUND OF INVENTION

[0002] Cell cultures are used today as a tool for the toxicological evaluation of substances. Bioreactors reproduce an environment favorable to the development and organization of cells, close to that of a tissue or an animal or human organ. Thanks to them, one can predict the reaction of an organ to a substance such as a xenobiotic, a cosmetic, a drug or more generally any active principle, and develop relevant in vitro models. These models are increasingly used in pharmaceutical research as they represent a serious alternative to in vivo models, z.e., animal experimentation, against which both economic and ethical pressures are emerging at the international level. Cell culture also represents the basis for the creation of artificial organs capable of replacing failing or absent organs, one of the challenges of tissue engineering. The bioreactors used in dynamic cell culture include at both ends of a culture chamber a fluid inlet and a fluid outlet to allow the passage of a nutritive fluid necessary for the development of cells. Various cell culture systems have been proposed for this purpose. However, these known systems are not satisfactory.

[0003] Indeed, the known culturing systems do not allow to take into account for the biological barrier (such as the skin or the stomach) interacting with the substance before reaching the organ in which the reaction with said substance is analyzed.

[0004] Some fluidic systems provide a barrier membrane which simulate the biological barrier. However, the barrier membrane is mainly adapted for the use in a single reactor and is not adapted for the multi-bioreactors which have been developed to increase the cell culture surface by parallelizing the bioreactors such as the multi-well plates.

[0005] There is thus a need for a multi-well system which allows to simulate the biological barrier and which is easy to use.

[0006] To this end, a purpose of this invention is to provide a device for simulating a biological barrier configured for interfacing at least one biochip. The device comprises at least one set of wells fluidically connected to a biochip. At least one of the wells of each set of wells comprises a holding structure configured to receive a barrier membrane therefore allowing to set a flow through the barrier membrane to the biochip so that to simulate the biological barrier by the barrier membrane.SUMMARY

[0007] This invention thus relates to a device for simulating a biological barrier configured for interfacing at least one biochip, said device comprising:A multi- well plate comprising at least one set of wells, each set of wells comprising a collecting well and a barrier well, each of the collecting well and the barrier well having an upper extremity and a lower extremity,A port plate comprising at least two ports, the port plate being configured to cover the multi-well plate so that the upper extremity of each of the collecting well and the barrier well receives at least one of the ports, and A connection plate comprising at least two connectors configured to be connected to an inlet and an outlet of a biochip, the connection plate being configured to be disposed under the multi-well plate so that the lower extremity of each of the collecting well and the barrier well receives at least one of the connectors. wherein the barrier well comprises a holding structure configured to receive a barrier membrane so that the barrier membrane separates said barrier wells into an upper chamber and a lower chamber, the port plate covering the upper chamber, the connection plate sustaining the lower chamber, and the lower chamber comprising a bypass channel.

[0008] Indeed, the device allows to simulate, in vitro, the coculture of at least two cell cultures wherein one of the cell culture acts like a barrier (filter) and is received in the holding structure whereas cells of the second cell culture are received in the biochip. The port plate allows a fluidic connection between the device and, for example, a fluid circuit or a sampling pipette.

[0009] According to an advantageous aspect of the invention, each set of wells further comprises a reservoir well fluidically connected to the upper extremity of the barrier well.

[0010] The reservoir well allows to contain, for example, a nutrient solution. The constant fluidic connection with the reservoir well therefore allows a continuous supply or nutrient while letting the two ports of the port plate free for sampling.

[0011] According to an advantageous aspect of the invention, the device further comprises a first sealing element positioned between the port plate and the multi-well plate and a second sealing element positioned between the multi-well plate and the connection plate.

[0012] The sealing element prevent leaks around the wells.

[0013] According to an advantageous aspect of the invention, the device further comprises a fastening element configured to maintain the port plate and the connection plate abutted against the multi- well plate.

[0014] The fastening element leads to a device which is easy to manipulate since all the plates are maintained together and prevents leaks.

[0015] According to an advantageous aspect of the invention, the barrier well has an upper segment and a lower segment, each of the upper segment and the lower segment having a section, the section of the upper segment being greater than the section of the lower segment so that the barrier well has a tapering, the holding structure comprising the tapering.

[0016] According to an advantageous aspect of the invention, the upper chamber has a first volume and the lower chamber has a second volume lower than the first volume.

[0017] This allows to expose the upper chamber to a small quantity of substances such as growth factors which are expensive. The lower chamber can be perfused via the bypass channel with a larger quantity of substance which may be less expensive (such as nutrients). This allows a reduction of costs when using the device.

[0018] The invention also relates to a system for simulating a biological barrier, said device comprising:The device described hereabove, at least one barrier membrane held by the holding structure of at least one of the barrier wells.

[0019] According to an advantageous aspect of the invention, biological cells are on the barrier membrane.

[0020] According to an advantageous aspect of the invention, the barrier membrane is configured for growing biological cells.

[0021] According to an advantageous aspect of the invention, the barrier well has an upper segment and a lower segment, each of the upper segment and the lower segment having a section, the section of the upper segment being greater than the section of the lower segment so that the barrier well has a tapering, the barrier membrane being bound to the bottom of a hollow body having a body section smaller than the section of the lower segment, the hollow body being topped by a collar having a collar section larger than the section of the lower segment and smaller than the section of the upper segment.

[0022] The invention also relates to method for culturing cells using the system for simulating a biological barrier described hereabove, the method comprising the steps of: a. Disposing at least one cell to be cultured in the at least one biochip, b. For each biochip, setting a flow from the port of the barrier well through the upper chamber and the barrier membrane, the flow being filtered by the barrier membrane, the filtered flow then flowing through the lowerchamber, the inlet of the biochip and the outlet of the biochip towards the collecting well.

[0023] According to an advantageous aspect of the invention, the method further comprises a step of setting a second flow from the bypass channel of the lower chamber of the barrier well to the collecting well successively through the inlet of the biochip and the outlet of the biochip.BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 shows a set of a collecting well (left) and a barrier well (right), said barrier well comprising a circular tapering as holding structure. The barrier well comprises here a bypass channel.

[0025] Figure 2 shows a barrier membrane suitable to be inserted in the barrier well of figure 1.

[0026] Figure 3 shows the barrier membrane of figure 2 inserted in the barrier well of figure 1.

[0027] Figure 4 shows a multi-well plate with sets of wells each comprising a barrier well and a collecting well.

[0028] Figure 5 shows a multi- well plate with sets of wells each comprising a reservoir well, a barrier well and a collecting well. Here, in each set of wells, the reservoir well is fluidically connected to the barrier well.

[0029] Figure 6 shows a top view of the device with biochips connected to the wells.

[0030] Figure 7 shows a bottom view of the device with biochips connected to the wells.

[0031] Figure 8 shows a transparency view of a set of wells with the ports and the connectors.

[0032] Figure 9 represents the flow set in the system comprising a biochip.

[0033] Figure 10 shows a top view of the device with a fastening means in the form of a lever vice.

[0034] Figure 11 shows a bottom view of the device with biochips connected to the well and a fastening means in the form of a lever vice.DEFINITIONS

[0035] In the present invention, the following terms have the following meanings:

[0036] “Biochip” refers to any chip allowing for cell culture. The biochip may be a bioreactor, a reactor, a flow chamber, a perfused chamber, a perfused tissue culture polystyrene (TCPS) cell culture box, a dynamic cell culture system. The biochip may be microfluidic or millifluidic.

[0037] “Lower” refers to the position of an element at lower altitude compared to another component when the device is in use.

[0038] “Nested” refers to the inclusion of a segment of a first channel - for instance a port - into a segment of a second channel - for instance a well. The section and the length / height of the nested segment are thus smaller than those of the nesting segment.

[0039] “Section of a segment” refers to the size of the surface of the segment perpendicular to its longitudinal axis.

[0040] “Upper” refers to the position of an element at higher altitude compared to another component when the device is in use.DETAILED DESCRIPTION

[0041] The following detailed description will be better understood when read in conjunction with the drawings. For the purpose of illustrating, the device is shown in the preferred embodiments. It should be understood, however that the application is not limited to the precise arrangements, structures, features, embodiments, and aspect shown. The drawings are not drawn to scale and are not intended to limit the scope of the claims to the embodiments depicted. Accordingly, it should be understood that where features mentioned in the appended claims are followed by reference signs, such signs are included solely for the purpose of enhancing the intelligibility of the claims and are in no way limiting on the scope of the claims.

[0042] This invention relates to a device 100 for simulating a biological barrier.

[0043] The device 100 represented, for example, in figure 8 comprises: a multi- well plate 30 comprising at least one set of wells, a port plate 20 comprising at least two ports 22, and a connection plate 40 comprising at least two connectors 44 configured to be connected to an inlet and an outlet of a biochip 80.

[0044] As shown in figure 8, each set of wells of the multi-well plate 30 comprises two wells (32, 34) respectively named collecting well 34 and barrier well 32. As shown in figures 1 and 3, each well (32, 34) is defined by a longitudinal axis 35 around which a wall extends. Each well (32, 34), better represented in figure 1, has two open extremities - an upper extremity and a lower extremity - transversal to the longitudinal axis 35, preferably perpendicular to the longitudinal axis 35. The axes of the collecting well 34 and barrier well 32 of one set of wells are preferably parallel, even more preferably, the axes of all wells (32, 34) of the multi-well plate 30 are parallel. The wells (32, 34) are preferably in a cylindrical form so that they present a circular section perpendicular to the longitudinal axis 35.

[0045] The volume of each well (32, 34) is ranging between 0.5 mL and 8 mL, preferably between 3 mL and 6 mL. If the well (32, 34) has a circular section, the diameter of this well (32, 34) is ranging from 5 to 25 mm, preferably from 10 to 20 mm.

[0046] The wells (32, 34) of a set of wells are linked by a well support 37 perpendicularly to the longitudinal axis 35 of each well (32, 34). Preferably, the well support 37 has a height corresponding to the height or length of the wells, the height / length being measured parallelly to the longitudinal axis 35. In this embodiment, the wells (32, 34) are embodied in the well support 37 and appear like holes in the well support 37 as shown in figure 4.

[0047] Advantageously, the multi-well plate 30 may comprise at least two set of wells (32, 34). This therefore allows to parallelize the bioreactors to simulate, for example, a plurality of organs and the interactions between them. In this embodiment, all the wells (32, 34) of the multi- well plate 30 may be linked by a unique well support 37 as in figure 4. The wells (32, 34) are preferably arranged in a 2:3 ratio matrix comprising 2 lines and 3 rows (or inversely 2 rows and 3 lines) of wells (32, 34) and their multiples such as 4 lines and 6 rows, 6 lines and 9 rows, 8 lines and 12 rows, etc. For example, the multi-well plate 30 may comprise 6, 12, 24, 96 or 384 wells (32, 34).

[0048] The barrier well 32, represented on the right-hand side of figure 1, comprises a holding structure 38 configured to receive a barrier membrane 64 so that the barrier membrane 64 separates said barrier wells 32 into an upper chamber 32a and a lower chamber 32b as shown in figure 3.

[0049] The lower chamber 32b comprises a bypass channel 39. The bypass channel 39 preferably takes the form of a channel (or hole) through the wall of the barrier well 32. The bypass channel 39 may comprise a port 39b in order to easily interface the barrier well 32 with a fluidic circuit. The port 39b may comprise a plug, or any other equivalent device making the channel easily opened or closed. In one embodiment, the port 39b of the bypass channel 39 is intrinsically comprised in the device 100 of the invention so that it cannot be removed from the wall of the barrier well 32. In an alternative embodiment, the port 39b of the bypass channel 39 is reversibly bound to the wall of the barrier well,for example by screwing. The reversible binding is advantageous because if the port is defective, it may be replaced without need to replace the whole multi-well plate 30. Advantageously, the port 39b is made in a flexible material, for example in polypropylene. Preferably, each port 39b is a standard port of 4 mm of diameter and 10- 32 UNF.

[0050] In an embodiment, the barrier membrane 64 is an integral part of the barrier well 32. In other words, it may be irreversibly bound to the wall of the barrier well 32.

[0051] In an alternative embodiment, the barrier membrane 64 is not comprised in the device 100 and may be reversibly inserted into the barrier well 32. The barrier membrane 64 thus takes the form of an insert.

[0052] For example, the barrier well 32 has an upper segment 32’ and a lower segment 32” characterized by two different sections. The section of the upper segment 32’ - z.e., the largest dimension measured perpendicularly to the longitudinal axis 35 - is greater than the section of the lower segment 32” so that the barrier well 32 has a tapering 38.

[0053] The barrier membrane 64 may be bound to the bottom of a hollow body 63, represented in figure 2. The hollow body 63 has a body section smaller than the section of the lower segment 32”. For example, if the hollow body 63 has a circular section, the diameter of the hollow body 63 is ranging from 5 to 25 mm, preferably from 10 to 20 mm. The hollow body 63 is topped by a collar 62 having a collar section larger than the section of the lower segment 32” and smaller than the section of the upper segment 32’. Alternatively, the hollow body 63 comprises at least two protrusions at the extremity opposite to the barrier membrane 64. The length of the protrusions is smaller than the difference of the sections between the lower segment 32” and the upper segment 32’.

[0054] The tapering 38 of the barrier well 32 is able to receive the collar 62 or the protrusions of the barrier membrane 64. The dimensions of the collar 62 or the protrusions are thus defined so that the barrier membrane 64 may be inserted through theupper segment 32’ and is resting on the tapering 38 without sliding through the lower segment 32”.

[0055] The device 100 wherein at least one barrier membrane 64 is inserted thus forms a system 200 for simulating a biological barrier. Indeed, the system 200 allows to simulate, in vitro, the cooperation of at least two cell cultures. The biological barriers is simulated by the barrier membrane 64 which acts like a filter. The barrier membrane 64 may be a membrane artificially reproducing a filtering, a cell filtering or a cell barrier or the barrier membrane 64 may be configured to receive a cell culture therefore naturally reproducing the filter or barrier function of the cultured cells. The system 200 comprises a culturing chamber configured to receive a cell culture, preferably, the second cell culture. In a first embodiment, the culturing chamber is the lower chamber 32b of the barrier well 32 comprising the barrier membrane 64. The culturing chamber is thus comprised in the device 100. In a second embodiment, the culturing chamber is the barrier well 32 of another set of wells, the lower chamber 32b of the barrier well 32 comprising the barrier membrane 64 being fluidically connected to the other set of wells. In a third embodiment, the culturing chamber is a biochip 80 which may be connected to the system 200, so that the inlet of said biochip 80 is connected to one connector 44 of the barrier well 32 comprising the barrier membrane 64 and the outlet of said biochip 80 is connected to another connector 44 of the collecting well 34. The barrier membrane 64 may be a layer of cells on a membrane. Preferably, the barrier membrane 64 is configured for growing biological cells. For example, the barrier membrane 64 may be structured in order to provide a tridimensional support optimizing the growth of the cells.

[0056] The port plate 20, shown in figure 6, covers the wells (32, 34). Preferably, the port plate 20 is abutted against the multi-well plate 30. The port plate 20 comprises at least two ports 22. More precisely, the port plate 20 is configured to cover the multi- well plate 30 so that the upper extremity (on the side of the upper segment 32’) of each well (32, 34) receives at least one of the ports 22. For example, as shown in figure 8, the upper segment 32’ of the barrier well 32 receives two ports 22 whereas the upper extremity of the collecting well 34 receives one port 22. The presence of more than one port 22 in a well (32, 34) is advantageous because it allows to let a continuous flowthrough one of the ports 22 of one well while having access into said well via the other port 22 in order to perform some material collection or analysis.

[0057] Each port 22 may comprise a plug, or any other equivalent device making the port 22 easily opened or closed.

[0058] In one embodiment, the ports 22 are intrinsically comprised in the port plate 20 so that they cannot be removed from the port plate 20. The port plate 20 may be manufactured by molding so that the ports 22 are made in a single piece with the port plate 20. For example, the material used is polystyrene plate, which is inexpensive and perfectly transparent, making it easy to observe the cells during culture. Each port 22 preferably extends along a longitudinal axis parallel to the longitudinal axes 35 of the wells (32, 34).

[0059] In an alternative embodiment, the ports 22 are reversibly bound to the port plate 20, for example by screwing. The reversible binding is advantageous because if one of the ports 22 is defective, this port may be replaced without need to replace the whole port plate 20. Advantageously, the ports 22 are made in a flexible material, for example in polypropylene so that they are less subject to damages. Preferably, the ports 22 are standard ports of 2 mm of diameter and 10-32 UNF.

[0060] The ports 22 may be configured to be nested in the wells (32, 34) so that at least part of the port is included into the well (32, 34). The section and the length of the nested part are thus smaller than the nesting well (32, 34).

[0061] The connection plate 40, shown in figure 7, is configured to be disposed under the multi-well plate 30 so that the lower extremity (on the side of the lower segment 32”) of each well 32 receives at least one of the connectors 44. The connection plate 40 is preferably abutted against the multi-well plate 30. The connection plate 40 thus sustains the lower chamber 32b.

[0062] The surface of the connection plate 40 may be flat so that cells may be cultured in the lower chamber 32b. The culturing chamber is thus comprised in the barrier well 32.In this embodiment, the connectors 44 comprise a part which is nested / which extends inside the barrier well 32 to avoid the cultured cells to pass through the connectors 44.

[0063] Alternatively, the surface of the connection plate 40 may comprise curved parts corresponding to the positions of the wells, when sustaining the multi-well plate 30. The curvature allows to eliminate meniscus problems and edge effects.

[0064] A biochip 80 may be connected to the wells (32, 34) of one set of wells so that the inlet of said biochip 80 is connected to the connector 44 of the barrier well 32 and the outlet of said biochip 80 is connected to the connector 44 of the collecting well 34.

[0065] The connectors 44 of the same set of wells are preferably positioned so that the distance between them corresponds to the distance between the inlet and the outlet of a biochip 80. For example, the distance between the connectors 44 of the same set of wells ranges from 10 to 40 mm, preferably between 20 and 30 mm.

[0066] In order to correspond to the standard size used in the bioreactor domain, the ports 22 and the connectors 44 preferably have an inner diameter of about 2 mm.

[0067] From higher altitude to lower altitude when in use, the device 100 is architecturally composed of the port plate 20, the multi-well plate 30, the connection plate 40 to which at least one biochip 80 may be connected - see figures 6 and 7. The device 100 therefore allows a fluid to flow from one of the ports 22 to one of the connectors 44 through the well connecting said port 22 and said connector 44.

[0068] As shown in figure 5, each set of wells may further comprise a reservoir well 34b fluidically connected to the upper segment 32’ of the barrier well 32.

[0069] As shown in figures 6 to 11, the device 100 may further comprise a first sealing element 60a positioned between the port plate 20 and the multi-well plate 30. The device 100 may further comprise a second sealing element 60b positioned between the multi-well plate 30 and the connection plate 40. In this embodiment wherein the device 100 comprises a sealing element (60a, 60b), the abutment between the port plate 20 and the multi-well plate 30 or the multi-well plate 30 and the connection plate 40corresponds to a contact between the plates (20, 30, 40) and the sealing element (60a, 60b) disposed between them.

[0070] As shown in figures 7, 8, 10 and 11, the device 100 may further comprise a fastening element 50 configured to maintain the port plate 20 or the connection plate 40 abutted against the multi- well plate 30.

[0071] For example, as shown in figures 7 and 8, the fastening element 50 comprises at least one screw screwed through the port plate 20 and the multi-well plate 30, the connection plate 40 and the multi-well plate 30, or the port plate 20, the multi-well plate 30 and the connection plate 40.

[0072] In another embodiment represented in figures 10 and 11, the fastening element 50 is configured to maintain the port plate 20, the multi-well plate 30 and the connection plate 40 abutted against each other from the outside of the device 100 contrarily to screws. For example, the fastening element 50 is a clamping element. More particularly, the fastening element 50 may be a lever vice configured to evenly distribute a pressure along the port plate 20 and the connection plate 40. To do so, the fastening element 50 comprises an upper pressing element 51 configured to be uniformly disposed on the port plate 20 and a lower pressing element 53 configured to be uniformly disposed on the connection plate 40, the multi-well plate 30 and optionally the sealing elements (60a, 60b) being disposed between the port plate 20 and the connection plate 40. The lever vice is configured to apply a force on each of the pressing elements (51, 53) so that the port plate 20, the multi-well plate 30 and the connection plate 40 are pressed towards each other. This is particularly advantageous in the embodiment wherein the device 100 comprises sealing elements (60a, 60b). The pressure may be evenly distributed if, for example, each of the pressing elements (51, 53) is disposed all along the circumference of the port plate 20 and the connection plate 40.

[0073] In order to culture the cells using the system 200 for simulating a biological barrier described above, the following steps may be performed: First, at least one cell to be cultured is seeded in the culturing chamber.

[0074] In the embodiment wherein the culturing chamber is a biochip 80, said cell is disposed in the biochip 80 using, for example, a pipette inserted through the inlet or the outlet. The biochip 80 is then connected to the device 100 via the connection plate 40 so that the inlet is connected to the connector 44 received by the barrier well 32 and the outlet is connected to the connector 44 received by the collecting well 34 of one set of well.

[0075] Secondly, if not included in the device 100, the barrier membrane 64 is disposed in the barrier well 32 so that the barrier membrane 64 is held by the holding structure 38.

[0076] The port plate 20, the multi- well plate 30 and the connection plate 40 are abutted against each other thereby forming the system 200 for simulating a biological barrier.

[0077] Then, a flow 90 is set from the port 22 of said barrier well 32 through the upper chamber 32a and the barrier membrane 64, the flow being filtered by the barrier membrane 64, the filtered flow then flowing through the lower chamber 32b towards the collecting well 34. The flow can be collected from the collecting well 34 in order to be analyzed or may be flown back to the upper chamber 32a of the barrier well 32 in order to create a cycling fluidic circulation.

[0078] In the embodiment wherein the culturing chamber is the lower chamber 32b of the barrier well 32 comprising the barrier membrane 64, the collecting well 34 receiving the flow 90 may be the collection well 34 of the same set of wells as said barrier well 32. Alternatively, the collecting well 34 receiving the flow 90 belongs to another set of wells.

[0079] In the embodiment wherein the culturing chamber is the barrier well 32 of another set of wells, connector 44 of the barrier well 32 comprising the barrier membrane 64 may be fluidically connected to the port 22 of a barrier well 32 belonging to another set of wells which is connected to a collecting well 34. The flow 90 thus flows through two barrier wells 32 before reaching the collecting well 34.

[0080] In the embodiment wherein the culturing chamber is a biochip 80 as represented in figure 9, the filtered flow enters the biochip 80 via the inlet. The flow passes throughthe biochip 80 in order to bring nutrients to the growing cells, for example. The flow then gets out of the biochip 80 via the outlet and arrives in the collecting well 34.

[0081] It is thus possible to increase the number of coculturing cells by successively fluidically connecting the set of wells according to one or several embodiments hereabove.

[0082] The method may further comprise a step of setting a second flow 91 from the bypass channel 39 of the lower chamber 32b of the barrier well 32 to the collecting well 34 successively through the inlet of the biochip 80 and the outlet of the biochip 80. The second flow is thus not filtered by the barrier membrane 64.EXAMPLES

[0083] The present invention is further illustrated by the following example.

[0084] The device 100 comprises a multi- well plate 30 with 24 wells (32, 34) arranged in a 2:3 ratio matrix. The device 100 thus comprises 2 rows of 6 barrier wells 32 and 2 rows of 6 collecting wells 34. Each row is distributed along the length of the device 100. The device has a total length of 133 mm and a width of 98 mm.

[0085] The barrier well 32 comprises a holding structure 38 in the form of a tapering configured to receive a barrier membrane 64 comprising a collar 62.

[0086] The lower chamber 32b of each barrier well 32 comprises a bypass channel 39.

[0087] The port plate 20 covering the multi- well plate 30 is made in polycarbonate. The ports 22 are made in polypropylene. The port plate 20 has a height of 10 mm. The ports 22 have a height of 18mm above the port plate 22. The inner diameter of the ports 22 is 2 mm. The center of the ports 22 are separated by 19 mm along the length of the device 100. The first rows of wells (32, 34) as well as the second rows of wells (32, 34) are separated by 19 mm. The two rows of collecting wells 34 are separated by 28 mm.

[0088] The connection plate 40 disposed under the multi-well plate 30 is made in polycarbonate. The connectors 44 are made in polypropylene. The connecting plate 40 has a height of 10 mm. The inner diameter of the connectors 44 is 3 mm. The distance between two connectors 44 of the same set of wells (32, 34) is 28.68 mm.

[0089] The multi-well plate 30 has a height of 20 mm. The barrier wells 32 and collecting wells 34 have an inner diameter of 16 mm. The centers of the wells (32, 34) are at the same position as the center of the ports 22. The barrier well 32 has a tapering 38 configured to receive an insert comprising the barrier membrane 64.

[0090] The sealing elements (60a, 60b) are disposed all along the surface of the multiwell plate 30 between the port plate 20 and the multi- well plate 30 and between the multiwell plate 30 and the connection plate 40. The sealing elements (60a, 60b) have a thickness of 4 mm.NUMERICAL REFERENCES20: port plate / / 22: ports of the port plate / / 30: multi-well plate / / 32: barrier well / / 32’: upper segment / / 32”: lower segment / / 32a: upper chamber / / 32b: lower chamber / / 34: collecting well / / 34b: reservoir well / / 35: longitudinal axis / / 37: well support / / 38: holding structure / / 39: bypass channel / / 39b: port of the bypass channel / / 40: connection plate / / 44: connectors / / 50: fastening element / / 51: upper pressing element / / 53: lower pressing element / / 60a: first sealing element / / 60b: second sealing element / / 62: collar / / 63: hollow body / / 64: barrier membrane / / 80: biochip / / 90: flow / / 91: second flow / / 100: device for simulating a biological barrier / / 200: system for simulating a biological barrier

Claims

CLAIMS1. A device (100) for simulating a biological barrier configured for interfacing at least one biochip, said device comprising:A multi-well plate (30) comprising at least one set of wells, each set of wells comprising a collecting well (34) and a barrier well (32), each of the collecting well (34) and the barrier well (32) having an upper extremity and a lower extremity,A port plate (20) comprising at least two ports (22), the port plate (20) being configured to cover the multi-well plate (30) so that the upper extremity of each of the collecting well (34) and the barrier well (32) receives at least one of the ports (22),A connection plate (40) comprising at least two connectors (44) configured to be connected to an inlet and an outlet of a biochip, the connection plate (40) being configured to be disposed under the multiwell plate (30) so that the lower extremity of each of the collecting well (34) and the barrier well (32) receives at least one of the connectors (44), wherein the barrier well (32) comprises a holding structure (38) configured to receive a barrier membrane (64) so that the barrier membrane (64) separates said barrier wells (32) into an upper chamber (32a) and a lower chamber (32b), the port plate (20) covering the upper chamber (32a), the connection plate (40) sustaining the lower chamber (32b), and the lower chamber (32b) comprising a bypass channel (39).

2. The device (100) according to any one of claim 1, wherein each set of wells further comprises a reservoir well (34b) fluidically connected to the upper extremity of the barrier well (32).

3. The device (100) according to claim 1 or 2 further comprising a first sealing element (60a) positioned between the port plate (20) and the multi-well plate (30) and a second sealing element (60b) positioned between the multi-well plate (30) and the connection plate (40).

4. The device (100) according to any one of claims 1 to 3 further comprising a fastening element (50) configured to maintain the port plate (20) and the connection plate (40) abutted against the multi- well plate (30).

5. The device (100) according to any one of claims 1 to 4, wherein the barrier well (32) has an upper segment (32’) and a lower segment (32”), each of the upper segment (32’) and the lower segment (32”) having a section, the section of the upper segment (32’) being greater than the section of the lower segment (32”) so that the barrier well (32) has a tapering, the holding structure (38) comprising the tapering.

6. The device (100) according to any one of claims 1 to 5, wherein the upper chamber (32a) has a first volume and the lower chamber (32b) has a second volume lower than the first volume.

7. A system (200) for simulating a biological barrier, said device comprising:The device (100) according to any of claims 1 to 6, at least one barrier membrane (64) held by the holding structure (38) of at least one of the barrier wells (32).

8. The system (200) according to claim 7, wherein biological cells are on the barrier membrane (64).

9. The system (200) according to claim 7 or 8, wherein the barrier membrane (64) is configured for growing biological cells.

10. The system (200) according to any one of claims 7 to 9, wherein the barrier well (32) has an upper segment (32’) and a lower segment (32”), each of the upper segment (32’) and the lower segment (32”) having a section, the section of the upper segment (32’) being greater than the section of the lower segment (32”) so that the barrier well (32) has a tapering, the barrier membrane (64) being bound to the bottom of a hollow body having a body section smaller than the section of the lower segment (32”), the hollow body being topped by a collar (62) having a collarsection (62) larger than the section of the lower segment (32”) and smaller than the section of the upper segment (32’).

11. A method for culturing cells using the system (200) for simulating a biological barrier according to any of claims 7 to 10, the method comprising the steps of: a. Disposing at least one cell to be cultured in the at least one biochip (80), b. For each biochip (80), setting a flow (90) from the port (22) of the barrier well (32) through the upper chamber (32a) and the barrier membrane (64), the flow being filtered by the barrier membrane (64), the filtered flow then flowing through the lower chamber (32b), the inlet of the biochip (80) and the outlet of the biochip (80) towards the collecting well (34).

12. The method for culturing cells according to claim 11, the method further comprising a step of setting a second flow (91) from the bypass channel (39) of the lower chamber (32b) of the barrier well (32) to the collecting well (34) successively through the inlet of the biochip (80) and the outlet of the biochip (80).

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