Device for filtering mesoscopic cellular objects in suspension

Abstract

The invention relates to a device (1) for filtering mesoscopic cellular objects in suspension (OCMS), comprising a body (2) having at least a first port (5) and a second port (6) and a filtering membrane (3) extending along a plane P in the body (2), thus defining two compartments, and able to prevent the passage of the OCMS from one compartment to the other, the first port (5) being intended for the injection and withdrawal of the OCMS immersed in a soaking solution and / or a rinsing solution, and the second port (6) being intended for the withdrawal and injection of the soaking solution and / or the rinsing solution. According to the invention, the injection into the first port (5) takes place in a direction D that is not substantially orthogonal to the plane P of the filtering membrane (3).

Description

[0001] DEVICE FOR FILTERING MESOSCOPIC CELLULAR OBJECTS IN SUSPENSION

[0002] Technical field

[0003] The present invention relates to the technical field of devices for filtering mesoscopic cellular objects in suspension.

[0004] Mesoscopic cellular objects in suspension, hereinafter referred to as OCMS, comprise a set of objects whose dimensions vary between 5 pm and 5 mm, which are in suspension and which comprise at least one eukaryotic cell, encapsulated or not. For example, OCMS comprise human cells in suspension, microtissues, spheroids, cell aggregates, embryoid bodies, encapsulated or not.

[0005] State of the art

[0006] In the above field, OCMS filtration modules are known comprising a body comprising a filter membrane intended to retain the OCMS but for example capable of allowing the solution in which the OCMS are bathed to pass through.

[0007] Several filtration techniques exist, in particular frontal filtration, where the OCMS are injected normally at the plane of the filter membrane. This technique can cause the accumulation and compaction of OCMS on the membrane which can prevent the proper filtration of the OCMS, damage them and clog the filter.

[0008] Another filtration technique is tangential flow filtration, where the OCMS in solution are injected through a tube porous to the solution but not to the OCMS, in which the solution is withdrawn through the tube wall, while the OCMS pass through the tube. This tangential flow filtration technique minimizes the risk of accumulation and compaction. The disadvantage of this technique is that it relies on filters that are complex to manufacture and it is difficult to completely rinse the OCMS. Indeed, there is always a residual proportion of soaking solution after rinsing due to the fact that the OCMS are not confined in a compartment and multiple rinsing steps are required to achieve the desired rinse solution concentration.

[0009] Another filtration technique is centrifugal filtration, which does not use a membrane but uses the difference in density of OCMS compared to the soaking / rinsing solution to separate OCMS and solution. This technique requires heavy equipment (centrifuge), is expensive and complex to set up.

[0010] The present invention aims to overcome these drawbacks.

[0011] Summary of the invention The invention therefore relates to a device for filtering suspended mesoscopic cellular objects (OCMS) comprising a body comprising at least a first port and a second port and a filter membrane extending along a plane P in the body thus defining two compartments and capable of preventing the passage of OCMS from one compartment to the other, the first port being intended for the injection and withdrawal of OCMS bathed in a soaking solution and / or a rinsing solution and the second port being intended for the withdrawal and injection of the soaking solution and / or the rinsing solution.

[0012] According to the invention, the injection into the first port is carried out in a direction D not substantially orthogonal to the plane P of the filter membrane.

[0013] Advantageously, the injection into the first port is a confined tangential of the OCMS and thus makes it possible not to create accumulation or damage to the OCMS.

[0014] A tangential flow or vortex in the filtration device reduces damage to the OCMS by keeping the OCMS moving.

[0015] In one embodiment, the body is designed such that the shortest path of an OCMS in the body between its injection and its withdrawal is a curved trajectory, passing through only one of the two compartments. Preferably, only one of the compartments is designed to receive the OCMS injected through the first port, both compartments being designed to have a soaking solution and / or a rinse solution passing through them.

[0016] The device for filtering suspended mesoscopic cellular objects (OCMS) comprises a body comprising a first port and a second port and a filter membrane extending along a plane P in the body, thus defining two compartments and capable of preventing the passage of OCMS from one compartment to the other, the first port being intended for the injection and withdrawal of OCMS bathed in a soaking solution and / or a rinsing solution and the second port being intended for the withdrawal and injection of the soaking solution and / or the rinsing solution, the first port being a single port in the first compartment.

[0017] OCMS also refers to cellular microcompartments.

[0018] In this example, the OCMS, preferably the cellular microcompartments, injected through the first port into a first compartment are stored in this first compartment until they are withdrawn through the first port.

[0019] The implementation of a single port in the first compartment of the filtration device according to the invention allows the injection and withdrawal of the OCMS bathed in a soaking solution through the same port, thus reducing the number of channels and the use of tubes and also making it possible to obtain a more compact filtration device. Advantageously, the use of a single port for the injection and withdrawal of the OCMS, preferably the cellular microcompartments, bathed in a soaking solution and / or the rinsing solution makes it possible to reduce the number of constituent elements of the filtration device, thus limiting the impact of the environment such as risks of external contamination at the ports.

[0020] According to a characteristic of the invention, the direction D is parallel to the plane P.

[0021] According to one embodiment of the invention, the body comprises an inner casing with edges having a rounded shape and the useful section of the filter membrane has the shape of a circle or an ellipse. The implementation of rounded edges makes it possible to reduce the nooks inside the body of the device, thus limiting the accumulation of OCMS in the nooks and making it possible to avoid the loss of OCMS trapped in the device during the OCMS recovery phases. This also makes it possible to limit the dead volume and therefore the volume necessary to rinse the OCMS.

[0022] In one embodiment of the invention, the body has a perforated shape or a cavity in its envelope. It may in particular be a toric shape, the envelope of which is a surface generated by a closed generating curve, in particular a circle or an ellipse, circulating on another closed directrix curve, in particular a circle. This cavity or hole formed in the envelope makes it possible to create or reinforce a circular circulation of the OCMS in the body, and therefore to avoid a phenomenon of clogging of the filtering device.

[0023] According to an exemplary embodiment of the invention, the body has a toric shape and the useful section of the filter membrane has the shape of a circle or an ellipse. Advantageously, the toric shape makes it possible to increase the swirl effect for the OCMS while significantly reducing the dead volume inside the body, thus limiting the volume necessary to rinse the OCMS.

[0024] According to a characteristic of the invention, any section perpendicular to the plane P of the compartment intended to receive the OCMS has a surface area strictly less than the useful surface area of ​​the filter membrane.

[0025] According to another characteristic of the invention, any section perpendicular to the plane P of the compartment intended to receive the OCMS has a surface area twice, five times and preferably ten times smaller than the useful surface area of ​​the filter membrane.

[0026] The tangential injection associated with such a difference in dimension makes it possible to inject the OCMS with a certain flow rate, promoting the vortex, while allowing the concentration of a certain number of OCMS inside the body of the filtration device and minimizing the dead volume. According to a characteristic of the invention, the injection into the second port is also carried out in a direction that is non-orthogonal, and preferably parallel, to the plane P of the filter membrane. The second tangential injection makes it possible to promote the vortex flows inside the body of the filtration device.

[0027] According to yet another feature of the invention, the soaking solution is a solution allowing cell culture. The OCMS are thus maintained in a favorable environment and are not degraded.

[0028] According to a characteristic of the invention, the soaking solution contains ions from the following list: calcium, iron, barium, strontium, barium, iron copper lead, aluminum, magnesium, preferably calcium, iron, barium. The addition of such ions is of great interest for the good performance of the OCMS.

[0029] According to another characteristic of the invention, the filter membrane comprises a microfabric covered with a fiber membrane. The implementation of such a filter membrane makes it possible to maintain control over the type of OCMS to be filtered, preferably over the type of cellular microcompartment to be filtered.

[0030] According to another embodiment of the invention, the filter membrane is capable of filtering cellular microcompartments having a largest dimension of between 120 and 700 micrometers, preferably between 200 pm and 400 pm, preferably between 200 pm and 350 pm, even more preferably 200 pm and 300 pm, in particular between 200 pm and 250 pm. Where appropriate, the filter membrane comprises porosities of less than 100 micrometers, or even less than 50 micrometers, or even less than 10 micrometers. Preferably, the filter membrane is capable of filtering cellular microcompartments having a largest dimension of less than 600 micrometers, the filter membrane comprises porosities of less than 100 micrometers.

[0031] For the purposes of the invention, the term “smallest dimension” of a microcompartment means the value of the smallest Feret diameter of said microcompartment.

[0032] By "Feret diameter" of a microcompartment according to the invention, we mean the distance "d" between two tangents to said microcompartment (or to said part), these two tangents being parallel, such that the entire projection of said microcompartment (or of said part) is between these two parallel tangents. A Ferret diameter of the internal part of the microcompartment is measured between two interfaces of the internal part and the external layer of the microcompartment, i.e. the distance "d" between two tangents to said internal part, these two tangents being parallel, such that the entire projection of said internal part is between these two parallel tangents. According to another characteristic of the invention, the device comprises two compartments having different volumes.Preferably, the compartment intended to receive OCMS, preferably cellular microcompartments, bathed in a soaking solution and / or a rinsing solution has a volume greater than the compartment intended to receive the soaking solution and / or the rinsing solution. The difference in volume advantageously makes it possible to improve the efficiency and the quantity of microcompartments harvested. Preferably, the compartment intended to receive cellular microcompartments bathed in a soaking solution and / or a rinsing solution has a volume at least 50% greater than the microcompartment intended to receive the soaking solution and / or the rinsing solution.

[0033] According to one embodiment of the invention, the filter membrane is capable of allowing other OCMS to pass through, preferably other cellular microcompartments, of a different type contained in the soaking solution. This characteristic in particular allows filtering of a certain type of OCMS, preferably cellular microcompartments.

[0034] According to one embodiment of the invention, the filter membrane is capable of allowing other OCMS of a different type contained in the soaking solution to pass through. This characteristic in particular allows filtering of a certain type of OCMS.

[0035] The invention also relates to a method for accumulating OCMS comprising the following steps:

[0036] - Injection of OCMS bathed in the soaking solution into a first port of an OCMS filtration device according to the invention, filtration of the OCMS by the filter membrane and recovery of a portion of the soaking solution by the second port,

[0037] - Injection of the soaking solution into the second port and recovery of the accumulated OCMS through the first port.

[0038] According to one embodiment of the invention, the OCMS accumulation process is carried out continuously. The OCMS can thus be collected as quickly as they are produced, thus minimizing the residence time in the soaking solution.

[0039] The invention also relates to a method for cleaning OCMS comprising the following steps:

[0040] - Injection of OCMS bathed in a soaking solution into a first port of an OCMS filtration device according to the invention, filtration of the OCMS by the filter membrane and recovery of the soaking solution and the rinsing solution by the second port,

[0041] - Injection of a rinse solution into the first given port intended to replace the soaking solution and recovery of both AA and rinse solutions through the second port. Advantageously, the tangential flow reduces damage to the OCMS by minimizing contact with the filter membrane during the cleaning process.

[0042] According to a characteristic of the invention, the cleaning method further comprises a step of injecting the rinsing solution into the second port and recovering the cleaned OCMS via the first port.

[0043] The invention also relates to a cell encapsulation system, the system comprising at least two containers, one of the containers comprising a cell solution and the other of the containers comprising a solution capable of gelling, an encapsulation device comprising a fluidic chip comprising main and secondary inlets each being connected to one of the containers via a distributor and capable of forming a concentric jet from the solutions supplied by the distributors, the encapsulation device being arranged so that the jet splits, at the outlet of the encapsulation device, into drops of which the outer layer is the solution capable of gelling and the core the cell solution, a gelling bath arranged downstream of the encapsulation device to collect the drops formed by the encapsulation device and arranged to cause gelling of the outer layer of each drop during its immersion in the bath,and a filtration device according to the invention.,

[0044] Advantageously, the gelling bath and a single-use closed collection system are connected to the first port of the filtration device, in particular by a flow valve.

[0045] In an exemplary embodiment of the invention, the encapsulation system according to the invention comprises a control unit arranged to control a solution displacement device for injecting OCMS bathing in the soaking solution of the gelling bath into the first compartment of the filtration device via the first port, the OCMS being stopped by the filter membrane and accumulated in a first compartment of the filtration device, and said soaking solution being evacuated from the filtration device, passing from the first compartment to the second compartment via the filter membrane and then via the second port. A waste container may be connected to the second port of the filtration device to receive the evacuated soaking solution.

[0046] Advantageously, the control unit is arranged to control a solution displacement device for injecting a soaking solution into the second compartment of the filtration device through the second port, said solution being discharged from the filtration device by passing from the second compartment to the first compartment via the filter membrane and then through the first port, recovering the OCMS accumulated in the first compartment during the previous step. A container of a soaking solution may be connected to the second port of the filtration device to inject the soaking solution. The control unit may be arranged to control the flow valve to direct the OCMS recovered by the soaking solution to the single-use closed collection system.

[0047] Alternatively or cumulatively, in particular prior to the step of recovering the accumulated OCMS, the control unit is arranged to control a solution displacement device to inject a rinsing solution into the first compartment of the filtration device via the first port, said rinsing solution being discharged from the filtration device, passing from the first compartment, to rinse the accumulated OCMS, to the second compartment via the filter membrane and then via the second port.

[0048] Of course, the various features, variants and embodiments of the invention may be combined with each other in various combinations to the extent that they are not incompatible or mutually exclusive.

[0049] In addition, various other characteristics of the invention emerge from the appended description given with reference to the drawings which illustrate non-limiting embodiments of the invention and where:

[0050] Brief descriptions of the Figures:

[0051] [Fig.l] is a perspective view of an exemplary embodiment of a filtration device according to the invention,

[0052] [Fig.2] is a schematic view of an exemplary embodiment according to the invention,

[0053] [Fig.3] is a perspective view of an exemplary embodiment of a filtration device according to the invention,

[0054] [Fig.4] is a half-sectional view of Figure 3 along section plane AA,

[0055] [Fig.5] is a half-sectional view of another exemplary embodiment of the invention, and

[0056] [Fig.6] is a schematic view of an exemplary embodiment with two filtration devices according to the invention.

[0057] Detailed description of the invention

[0058] It should be noted that in these figures the structural and / or functional elements common to the different variants may have the same references.

[0059] The invention aims to filter suspended mesoscopic cellular objects, known as OCMS.

[0060] For these purposes, the invention designated by the reference 1 as a whole and illustrated in FIG. 1 relates to a filtration device comprising a body 2 comprising a filter membrane 3 which extends in the body 2 along a plane P.

[0061] According to the invention, the body 2 of the filtration device 1 comprises an inner casing 4 having rounded edges. According to one embodiment of the invention, particularly illustrated in Figures 2 and 5, the inner casing 4 has the shape of a burger, i.e. a cylindrical shape with a rounded upper edge surface and a rounded lower edge surface.

[0062] According to the embodiment illustrated in Figures 3 and 4, the inner envelope 3 of the body 2 has the shape of a donut, that is to say a toric shape.

[0063] According to the embodiment illustrated in Figure 2, the filtration device 1 is symmetrical with respect to the filter membrane 3. Other non-symmetrical embodiments illustrated in Figures 3, 4 and 5 are also compatible with the invention. For example, the part intended to receive OCMS may comprise larger volume dimensions than the part on the other side of the filter membrane 3 or vice versa.

[0064] The filter membrane 3 extends along a plane P in the body 2 of the filtration device 1. The filter membrane 3 extends to the edges of the inner casing 4 of the body 2 of the filtration device 1. According to the illustrated embodiments, the useful section of the filter membrane 3 has the shape of a circle. According to other embodiments not illustrated, the useful section of the filter membrane 3 has the shape of an ellipse.

[0065] In order to retain the OCMS, the filter membrane 3 comprises a fiber membrane capable of preventing the passage of the OCMS.

[0066] The filter membrane 3 is therefore adapted according to the type of OCMS implemented.

[0067] Suspended mesoscopic cellular objects (MSCOs) refer to single cells or microtissues, but also to cells encapsulated in alginate microcompartments.

[0068] OCMS are immersed in a soaking solution. Advantageously, the soaking solution allows for cell culture. For example, OCMS are cells encapsulated in alginate microcompartments and the soaking solution contains Calcium Chloride CaCl2.

[0069] According to one embodiment of the invention, the filter membrane 3 comprises a microfabric covered with a fiber membrane. The fiber membrane contains a material of the wide mesh textile type (mesh), nylon, or PET. Other filter materials, OCMS, are used within the scope of the invention.

[0070] According to one embodiment of the invention, for OCSMS, and more particularly for cellular microcompartments, having a largest dimension of between 120 and 700 micrometers, the filter membrane comprises porosities of less than 100 micrometers. According to one embodiment of the invention, for OCSMS, and more particularly for cellular microcompartments, having a largest dimension of less than 600 micrometers, the filter membrane comprises porosities of less than 100 micrometers.

[0071] According to one embodiment of the invention, the filter membrane 3 is capable of filtering at least one type of OCMS and is also capable of allowing another type of OCMS to pass through. The mesh can thus be adapted according to the dimensions of the OCMS to be filtered.

[0072] The body 2 of the filtration device 1 comprises a first port 5 intended for tangential injection of the OCMS bathed in a soaking solution.

[0073] The first port 5 comprises for its purposes a cylindrical shape which extends substantially in a direction D belonging to a plane P' parallel to the plane P of the filter membrane 3 as shown in figure 1.

[0074] The second port 6 is located in the second part of the body 2, on the other side of the filter membrane 3. According to the embodiment illustrated in figures 1 and 2, the second port 6 does not allow tangential injection into the body 2 of the filtration device 1. According to the embodiment illustrated in particular in figures 3, 4 and 5, the second port 6 is also suitable for tangential injection into the body 2 of the filtration device 1.

[0075] According to the embodiment illustrated in Figure 1, the first port 5 comprises an inlet 7 and an injection channel 8.

[0076] The injection channel 8 allows tangential injection into the inner casing 4 of the body 2 of the filtration device 1.

[0077] According to other embodiments not illustrated, the body 2 of the filtration device 1 comprises more than two inlet and outlet ports thus allowing the injection of different types of solution or OCMS on each side of the filter membrane 3.

[0078] The filtration device 1 according to the invention is used for different functions.

[0079] The filtration device 1 is used in particular for various processes such as OCSM accumulation and OCMS cleaning.

[0080] A first use of the filtration device 1 relates to an OCMS accumulation process.

[0081] A first step of the OCMS accumulation method consists of injecting OCMS bathed in the soaking solution into the first port 5 of the OCMS filtration device 1 according to the invention. The injection is carried out at a flow rate Q1 for a time T1. For example, the flow rate Q1 is 130 mL / min and the time T1 is 45 s. The OCMS are stopped by the filter membrane 3. A portion of the soaking solution is discharged through the second port 6 of the filtration device 1. A second step of the OCMS accumulation method consists of reversing the flow. To do this, the previously discharged soaking solution is injected into the second port 6.

[0082] The OCMS accumulated in the filtration device 1 during the first stage of the OCMS accumulation process are then recovered via the first port 5.

[0083] Such a method of accumulating OCMS using the filtration device 1 according to the invention can advantageously be continuous by repeating the two steps described.

[0084] A second use of the filtration device 1 according to the invention relates to a method for cleaning OCMS.

[0085] A first step of the OCMS cleaning process consists of injecting OCMS bathed in a soaking solution into the first port 5 of the filtration device 1 at a flow rate Q2, and filtering the OCMS by the filter membrane 3 and recovering a portion of the soaking solution through the second port 6 for a time T2. For example, the flow rate Q2 is 260 mL / min and the time T2 is 15 s.

[0086] A second step of the OCMS cleaning process consists of injecting a rinsing solution into the first port 5 at a flow rate Q3 for a given time T3, the rinsing solution then replacing the soaking solution and recovering the soaking solution mixed with the rinsing solution and then the rinsing solution alone via the second port 6. For example, the flow rate Q3 is 250 ml / min and the time T3 is 20 s.

[0087] A third step consists of reversing the flow with the injection into the second port 6 of the rinsing solution at a flow rate Q4 and the recovery of the cleaned OCMS by the first port 5 for a time T4. For example, the flow rate Q4 is 250 mL / min and the time T4 is 10 s.

[0088] Such an OCMS cleaning method using the filtration device 1 according to the invention can advantageously be continuous by repeating the three steps described.

[0089] Figure 6 illustrates the implementation of the two processes described above in series. A first filtration device 1 is used for an OCMS accumulation process and a second filtration device is used for an OCMS cleaning process.

[0090] Two filtration devices 1 are used to accumulate the produced OCMS and transfer them, and to wash the OCMS with a rinsing solution and to collect the OCMS in a closed single-use system. To do this, a flow valve 10 is interposed between the first two respective ports of the two filtration devices 1. And the steps described above can take place continuously.

[0091] The first port 5 of a first filtration device 1 is connected to the first port 5 of a second filtration device 1 in order to be able to directly inject OCMS accumulated by the first filtration device 1 and to clean them using the second filtration device 1.

[0092] Other architectural forms may be shaped according to the use of the invention.

[0093] Of course, various other modifications may be made to the invention within the scope of the appended claims.

Claims

CLAIMS

1. Filtration device (1) for suspended mesoscopic cellular objects (OCMS) comprising a body (2) comprising at least a first port (5) and a second port (6) and a filter membrane (3) extending along a plane P in the body (2) thus defining two compartments and capable of preventing the passage of the OCMS from one compartment to the other, the first port (5) being intended for the injection and withdrawal of the OCMS bathed in a soaking solution and / or a rinsing solution and the second port (6) being intended for the withdrawal and injection of the soaking solution and / or the rinsing solution, characterized in that the injection into the first port (5) is carried out along a direction D not substantially orthogonal to the plane P of the filter membrane (3).

2. Filtration device (1) according to the preceding claim in which the direction D is parallel to the plane P.

3. Filtration device (1) according to one of the preceding claims in which the body (2) comprises an inner casing with edges having a rounded shape and the useful section of the filter membrane (3) has the shape of a circle or an ellipse.

4. Filtration device (1) according to one of the preceding claims in which any section perpendicular to the plane P of the compartment intended to receive the OCMS has a surface area strictly less than the useful surface area of ​​the filter membrane (3).

5. Filtration device (1) according to the preceding claim in which any section perpendicular to the plane P of the compartment intended to receive the OCMS has a surface area twice, five times and preferably ten times smaller than the useful surface area of ​​the filter membrane.

6. Filtration device (1) according to one of the preceding claims in which the body (2) has a toric shape and the useful section of the filter membrane (3) has the shape of a circle or an ellipse.

7. Filtration device (1) according to one of the preceding claims in which the injection into the second port (6) is also carried out in a direction which is non-orthogonal, and preferably parallel, to the plane P of the filter membrane (3).

8. Filtration device (1) according to one of the preceding claims in which the soaking solution is a solution allowing cell culture.

9. Filtration device (1) according to the preceding claim in which the soaking solution contains ions from the following list: calcium, iron, barium, strontium, barium, iron copper lead, aluminum, magnesium, preferably calcium, iron, barium.

10. Filtration device (1) according to one of the preceding claims in which the filter membrane (3) comprises a microfabric covered with a fibrous membrane.

11. Filtration device (1) according to one of the preceding claims in which the filter membrane (3) is capable of allowing other OCMS of a different type contained in the soaking solution to pass through.

12. A method of accumulating OCMS comprising the following steps: - Injection of OCMS bathed in the soaking solution into a first port (5) of a filtration device (1) according to one of claims 1 to 11, filtration of the OCMS by the filter membrane (3) and recovery of a portion of the soaking solution by the second port (6), - Injection of the soaking solution into the second port (6) and recovery of the accumulated OCMS through the first port (5).

13. A method of cleaning OCMS comprising the following steps: - Injection of OCMS bathed in a soaking solution into a first port (5) of a filtration device (1) according to one of claims 1 to 11, filtration of the OCMS by the filter membrane (3) and recovery of the soaking solution and the rinsing solution by the second port (6), - Injection of a rinsing solution into the first port (5) given to replace the soaking solution and recovery of the two AA and rinsing solutions through the second port (6).

14. A method of cleaning OCMS according to the preceding claim further comprising a step of injecting the rinsing solution into the second port (6) and recovering the cleaned OCMS through the first port (5).

15. A cell encapsulation system, the system comprising at least: - two containers, one of the containers containing a cell solution and the other of the containers containing a solution capable of gelling, - an encapsulation device comprising a fluidic chip having main and secondary inlets each connected to one of the containers via a distributor and capable of forming a concentric jet from the solutions supplied by the distributors, the encapsulation device being arranged so that the jet splits, at the outlet of the encapsulation device, into drops of which the outer layer is the solution capable of gelling and the core the cell solution, - a gelling bath arranged downstream of the encapsulation device to collect the drops formed by the encapsulation device and arranged to cause gelling of the outer layer of each drop when it is immersed in the bath, and - a filtration device (1) according to one of claims 1 to 11.

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