Microfluidic system and method for processing a sample using said system

Abstract

The invention relates to a microfluidic cartridge (1) comprising a support in which a microfluidic circuit is formed, the microfluidic circuit comprising: A filtration chamber (10) in which a filter (F) is placed, said filter (F) dividing said filtration chamber (10) into a first space (100) positioned below the filter (F) and a second space (101) positioned above the filter (F), A first reservoir (R1) for receiving a sample, A first microfluidic channel (C1) connected on one side to the first reservoir (R1) and opening on the other side into the first space (100) of the filtration chamber (10), A second reservoir (R2), A second microfluidic channel (C2) connected on one side to the second reservoir (R2) and opening on the other side into the first space (100) of the filtration chamber (10). Figure to be published with the abbreviation: Figure 1

Description

Title of the invention: Microfluidic system and method for processing a sample using said system Technical field of the invention

[0001] The invention relates to a microfluidic cartridge, a microfluidic system integrating said microfluidic cartridge and a method for processing a sample implemented using said microfluidic system. State of the art

[0002] Microfluidic cartridges, also called microfluidic cards or microfluidic chips, incorporating a microfluidic circuit are well known in the prior art. They can be used to prepare a sample, containing biological species, before sample analysis. In particular, they can be used to filter the biological species present in the sample. To perform this filtration, the cartridge incorporates a filter adapted to trap the target biological species present in the sample.

[0003] When the sample passes through the filter, it very frequently becomes blocked, particularly due to the presence of bubbles on its surface. In other words, it does not perform its function, or it is necessary to apply strong pressure to force the sample through the filter. However, the presence of bubbles is often unavoidable, due in particular to the presence of air in the dead volumes of the microfluidic circuit or the nature of the buffers used (detergent).

[0004] A first objective of the invention is to provide a microfluidic cartridge having an architecture adapted to allow unblocking of the filter. Description of the invention

[0005] This objective is achieved by a microfluidic cartridge comprising a support in which a microfluidic circuit is formed, the microfluidic circuit comprising: - A filtration chamber in which a filter is placed, said filter separating said filtration chamber into a first space positioned below the filter and a second space positioned above the filter, - A first reservoir intended to receive a sample, - A first microfluidic channel connected on one side to the first reservoir and opening on the other side into the first space of the filtration chamber, and a first valve arranged on the first microfluidic channel, - A second reservoir, - A second microfluidic channel connected on one side to the second reservoir and opening on the other side into the first space of the filtration chamber, - A second valve arranged on the second microfluidic channel, - A third microfluidic channel opening into the second space (of the filtration chamber), and a third valve arranged on the third microfluidic channel, - The first tank and the second tank each comprising a chamber and a deformable membrane in the chamber to be able to move a fluid.

[0006] According to one particular feature, the microfluidic circuit comprises: - A third tank, - A fourth microfluidic channel connected on one side to said third reservoir and opening on the other side into the second space of the filtration chamber, said third reservoir also comprising a chamber and a deformable membrane in said chamber, - A fourth valve arranged on the fourth microfluidic channel.

[0007] Another object of the invention is to propose a microfluidic system adapted to unblocking the filter and a method for implementing the unblocking of the filter.

[0008] The invention therefore also relates to a microfluidic system comprising a microfluidic cartridge, actuation means and a control unit for said actuation means, the microfluidic cartridge being as defined above, the actuation means being arranged and configured to control the displacement of each membrane and the state of each valve of the microfluidic circuit.

[0009] The invention also relates to a method for treating a fluidic sample comprising biological species, implemented in a microfluidic system as defined above, said method comprising: - A first control step upon opening the first and second valves and closing the third valve, - A second step of injecting the sample from the first reservoir to the second reservoir through the first space of the filtration chamber, - A third step of injecting the sample from the second reservoir into the first reservoir through the first space of the filtration chamber, - A fourth control step involving the closing of the second valve and the opening of the third valve, - A fifth step involves injecting the sample from the first reservoir into the third microfluidic channel, passing through the first space of the filtration chamber, the filter, and the second space of the chamber. filtration, so as to trap said biological species in the first space of the filtration chamber.

[0010] According to one particular feature, a cycle comprising the first control step, the second injection step and the third injection step is implemented several times, prior to the execution of the fourth control step.

[0011] According to another feature, the process includes a sixth step of lysis of the biological species trapped in the first space of the filtration chamber, in order to release biological material.

[0012] According to another feature, said process also comprises: - A seventh control step at the closing of the third valve and the opening of the fourth valve, - An eighth step involving the injection of an eluting fluid from the third tank to the first tank or from the first tank to the third tank, making a round trip, - A ninth step of elution of the biological material by injection of the elution fluid from the first reservoir to the third reservoir passing through the first space of the filtration chamber, the filter and the second space of the filtration chamber. Brief description of the figures

[0013] Other features and advantages will become apparent in the detailed description that follows, given in relation to the accompanying drawings in which: - Figure 1 schematically illustrates the principle of implementation of the microfluidic system of the invention; - Figure 2 shows the microfluidic cartridge of the invention for the purpose of above ; - Figures 3A and 3B illustrate the operating principle of a microfluidic capsule; - Figures 4A to 4C illustrate the different stages of the treatment process according to the invention; - Figure 5 shows an example of a multi-layered cartridge architecture microfluidics of the invention;

[0014] Detailed description of at least one embodiment

[0015] With reference to [Fig. 1] and [Fig. 2], the microfluidic system comprises: - A microfluidic cartridge 1 in which a microfluidic circuit is made; - Means of actuation 2 of the microfluidic elements present in the microfluidic circuit, advantageously of pneumatic type; - A control unit UC intended to control the actuation means 2;

[0016] The microfluidic cartridge 1 is in the form of a support made of a transparent material, for example COC, PMMA or equivalent.

[0017] The support has an extensive surface area in two dimensions and a low thickness with respect to these two dimensions.

[0018] The support is made of several layers sealed together. An example of the architecture will be described below in relation to [Fig. 5]. The microfluidic circuit is integrated into said support by engraving / machining / embossing (or other known techniques) performed on one or more layers of the support. The same applies to the (pneumatic) control circuit used for controlling the fluidic elements of the cartridge.

[0019] The microfluidic circuit of a microfluidic cartridge comprises several fluidic elements, including several reservoirs and several valves.

[0020] With reference to [Fig. 3A] and [Fig. 3B], each reservoir is in the form of a microfluidic capsule having a chamber 3 in which a deformable membrane 30 is integrated. The membrane 30 is arranged so as to separate the chamber into a first volume belonging to the microfluidic circuit and a second volume belonging to the control circuit. By modulating the air pressure in the second volume, the displacement of the membrane 30 within the chamber is controlled, and thus the volume of fluid present in the first volume of the chamber. Depending on the direction of movement of the membrane, the fluid 31 is drawn into the first volume of the chamber (application of a negative pressure (P-) - [Fig. 3A]) or pushed out of the first volume of the chamber (application of a positive pressure (P+) - [Fig. 3B]).

[0021] Each valve is also advantageously of the pneumatic type, controllable between an open state in which it allows the fluid to pass through and a closed state in which it blocks the passage of fluid.

[0022] In the context of the invention, the microfluidic circuit comprises the following: - A filtration chamber 10. This filtration chamber 10 includes a filter F separating the chamber into a first space 100 located below the filter and a second space 101 located above the filter. The terms "below" and "above" are of course to be considered in a non-limiting manner and are to be understood when the cartridge is laid flat. - A first RI reservoir intended to receive a sample. - A first microfluidic channel Cl connected on one side to the first reservoir RI and opening on the other side into the first space 100 of the filtration chamber 10. - A second R2 purge tank. - A second microfluidic channel C2 connected on one side to the second purge reservoir R2 and opening on the other side into the first space 100 of the filtration chamber 10. - A third microfluidic channel C3 opening into the second space 101 of the filtration chamber 10. - A microfluidic outlet OUT through which the sample can be evacuated via the third microfluidic channel C3.

[0023] The filter F is in the form of a grid, having a mesh, the size of whose pores is adapted to the size of the biological species E to be trapped.

[0024] Within the framework of the invention, the first tank RI and the second tank R2 have the architecture already described above.

[0025] In addition, the microfluidic circuit incorporates: - A first valve V1 arranged on the first microfluidic channel Cl to control the passage of fluid through this first channel; - A second valve V2 arranged on the second microfluidic channel C2 to control the passage of fluid through this second channel; - A third valve V3 arranged on the third microfluidic channel C3 to control the passage of fluid through this third channel;

[0026] In addition, the microfluidic circuit can also integrate another reservoir (designated R4) intended to receive an elution fluid and a fourth microfluidic channel C4 connecting this reservoir R4 to the second space 101 of the filtration chamber 10. A fourth valve V4 is also arranged on this fourth microfluidic channel C4 to control the passage of fluid through this fourth channel C4.

[0027] This R4 tank has an architecture identical to that of the RI and R2 tanks described above.

[0028] Each RI, R2 and R4 reservoir of the microfluidic circuit may have several inlets / outlets accessible from the outside allowing the reservoir to be supplied with distinct compounds or a compound to be removed from the reservoir.

[0029] As indicated above, the system integrates the actuation means 2, advantageously pneumatic, used to control each valve in the open or closed state and the movement of fluid in each tank, by actuation of the diaphragm.

[0030] It should be noted that the microfluidic cartridge can incorporate, within its support, a single layer forming a membrane common to all the actuated fluidic elements (valve, reservoir in particular). By applying a localized positive or negative pressure to said membrane, the fluidic element is actuated. corresponding. This architecture is described in particular in relation to [Fig.5] below.

[0031] The system also incorporates a control unit UC responsible for controlling the actuation means 2 to execute a sequence. The sequence comprises several steps carried out successively, each step consisting of controlling one or more fluidic elements of the fluidic circuit to enable the implementation of the treatment process of the invention.

[0032] Based on the system architecture described above, in conjunction with Figures 4A to 4C, the processing method of the invention is as follows:

[0033] The sample is initially placed in the first RI reservoir, this sample containing biological species E.

[0034] El - [Fig.4A]: The actuation means 2 are controlled to open the first valve VI and the second valve V2, and close the third valve V3.

[0035] E2 - [Fig.4A]: The actuation means 2 are controlled to allow injection of the sample from the first reservoir RI to the second reservoir R2 through the first space 100 of the filtration chamber 10. As the third valve V3 is closed, the sample does not pass through the filter F. The sample is thus transferred into the second reservoir R2.

[0036] E3 - [Fig. 4A]: The actuation means 2 are controlled to allow the sample to be injected from the second reservoir R2 to the first reservoir RI through the first opening 100 of the filtration chamber 10. The reverse process is then repeated to return the sample to the first reservoir RL. This round-trip cycle can be performed once or several times. Its purpose is to clean the filter F of any bubbles that may be present on its surface and to unblock it, in preparation for the filtration of the sample.

[0037] E4 - [Fig.4B]: The actuation means 2 are controlled to close the second valve V2 and open the third valve V3. The first valve VI remains in the open state.

[0038] E5 - [Fig.4B]: The actuation means 2 are controlled for the injection of the sample from the first reservoir RI to the third microfluidic channel C3, passing through the first space 100 of the filtration chamber 10, the filter F and the second space 101 of the filtration chamber 10, so as to trap said biological species E in the first space 100 of the filtration chamber 10.

[0039] Before any filtration of a sample or any other compound, steps E1 to E3 can advantageously be implemented to unblock the filter and improve subsequent filtration. Thanks to the invention, complex analytical protocols using multiple samples and reagents can thus be performed. In particular, it is possible to use a washing liquid which is passed through the first space of the filtration chamber, between the RI and R2 tanks to wash and unblock the F filter.

[0040] Furthermore, thanks to the architecture of the system of the invention, it is also possible to foresee the following additional steps:

[0041] E6 - [Fig.4B]: It is possible to carry out a lysis (chemical and / or mechanical) of the biological species E trapped in the first space of the filtration chamber, in order to release biological material M (DNA for example).

[0042] E7 - [Fig.4C]: The actuation means 2 are controlled to close the third valve V3 and open the fourth valve V2. The first valve remains in the open state.

[0043] E8 - [Fig. 4C]: The actuation means are controlled for injecting an elution fluid from reservoir R4 to the first reservoir RI, through the fourth microfluidic channel C4, the second space 101 of the filtration chamber 10, the filter F, the first space 100 of the filtration chamber 10, and the first microfluidic channel CL. It would also be possible to inject the elution fluid from reservoir RI to reservoir R4 and make several round trips until the elution fluid is recovered from reservoir RL.

[0044] E9 - [Fig. 4C]: The actuation means 2 are controlled for injecting the elution fluid from the first reservoir RI to the reservoir R4, passing through the first microfluidic channel C1, the first space 100 of the filtration chamber 10, the filter F, the second space 101 of the filtration chamber 10, and the fourth microfluidic channel C4. The elution fluid thus carries the biological material M through the filter F to the reservoir R4. It would also be possible to provide for the elution of the biological material to the fluidic outlet OUT or to any other available outlet point above the filter F.

[0045] It should be noted that it would be possible to separate the volume recovered in the R4 tank into several fragments, each fragment being collected in a separate tank.

[0046] Figure 5 shows an example of an embodiment of the microfluidic cartridge 1. A microfluidic cartridge thus comprises several superimposed layers assembled together by sealing, adhesion, or other known techniques. The cartridge may therefore include: - A first lower layer L1 (the lowest) in which the first space 100 of the filtration chamber 10 is made. - On this first layer L1, possibly using an adhesive layer (not shown) with perforations in certain areas, a second layer L2 is assembled, in which the second space 101 is created. the filtration chamber. This second layer L2 may also include several pneumatic actuation inlets. - On the second layer L2, a third layer L3 is added, forming the membrane. This is a so-called full-plate membrane (perforated in a judicious way - particularly at the level of the filter F), meaning that it is common to all the pneumatically actuated fluidic elements of the cartridge 1. - On the third layer L3, a fourth layer L4 is added carrying the fluidic elements of the microfluidic circuit that can be acted upon by the membrane, that is to say the reservoirs RI, R2, R4 and the valves VI, V2, V3, V4.

[0047] It should be noted that: - The filter F is positioned between the first layer L1 and the second layer L2, to separate the filtration chamber 10 into its two spaces 100, 101 made respectively in the first layer L1 and the second layer L2. - Fluidic passages are provided through the layers of the cartridge to connect the fluidic elements of the fourth layer L4 to the two spaces 100, 101 of the filtration chamber 10. - The membrane is perforated in certain places, particularly at the filter, to allow the fluid to pass through the filter F.

[0048] The invention thus offers numerous advantages, including: - A simplified solution to unblock a filter F of a microfluidic cartridge 1; - A simple and reliable method for unblocking the F filter;

Claims

Demands

1. Microfluidic cartridge (1) comprising a support in which a microfluidic circuit is implemented, the microfluidic circuit comprising: - A filtration chamber (10) in which a filter (F) is placed, said filter (F) separating said filtration chamber (10) into a first space (100) positioned below the filter (F) and a second space (101) positioned above the filter (F), - A first reservoir (RI) intended to receive a sample, - A first microfluidic channel (Cl) connected on one side to the first reservoir (RI) and opening on the other side into the first space (100) of the filtration chamber (10), and a first valve (VI) arranged on the first microfluidic channel (Cl), - Characterized by the fact that the microfluidic circuit comprises: - A second tank (R2), - A second microfluidic channel (C2) connected on one side to the second reservoir (R2) and opening on the other side into the first space (100) of the filtration chamber (10), - A second valve (V2) arranged on the second microfluidic channel (C2), - A third microfluidic channel (C3) opening into the second space (101) of the filtration chamber (10), and a third valve (V3) arranged on the third microfluidic channel (C3), - The first reservoir (RI) and the second reservoir (R2) each comprising a chamber and a deformable membrane in the chamber to be able to move a fluid.

2. Microfluidic cartridge according to claim 1, characterized in that the microfluidic circuit comprises: - A third tank (R4),

3.

4. - A fourth microfluidic channel (C4) connected on one side to said third reservoir (R4) and opening on the other side into the second space (101) of the filtration chamber, said third reservoir (R4) also comprising a chamber and a deformable membrane in said chamber, - A fourth valve (V4) arranged on the fourth microfluidic channel (C4). Microfluidic system comprising a microfluidic cartridge, actuation means (2) and a control unit (CU) of said actuation means (2), characterized in that the microfluidic cartridge (1) is as defined in claim 1 or 2, and in that the actuation means (2) are arranged and configured to control the displacement of each membrane and the state of each valve of the microfluidic circuit. A method for treating a fluidic sample comprising biological species (E), implemented in a microfluidic system as defined in claim 3, said method being characterized in that it comprises: - A first control step (El) triggers the opening of the first valve (VI) and the second valve (V2) and the closing of the third valve (V3), - A second step (E2) of injecting the sample from the first reservoir (RI) to the second reservoir (R2) through the first space (100) of the filtration chamber, - A third step (E3) of injecting the sample from the second reservoir (R2) into the first reservoir (RI) through the first space (100) of the filtration chamber, - A fourth control step (E4) triggers the closing of the second valve (V2) and the opening of the third valve (V3), - A fifth step (E5) of injecting the sample from the first reservoir (RI) into the third microfluidic channel (C3), passing through the first space (100) of the filtration chamber, the filter (F) and the second space (101) of the filtration chamber, so as to trap said biological species (E) in the first space (100) of the filtration chamber.

5. Method according to claim 4, characterized in that a cycle comprising the first control step (E1), the second injection step (E2) and the third injection step (E3) is carried out several times, prior to the execution of the fourth control step (E4).

6. A method according to claim 4 or 5, characterized in that it comprises a sixth step (E6) of lysis of the biological species (E) trapped in the first space (10) of the filtration chamber, in order to release biological material (M).

7. A method according to claim 6, characterized in that the microfluidic system comprises a microfluidic cartridge (1) as defined in claim 2, said method comprising: - A seventh step (E7) of controlling the closing of the third valve (V3) and the opening of the fourth valve (V4), - An eighth step (E8) of injecting an elution fluid from the third reservoir (R4) to the first reservoir (RI) or from the first reservoir (RI) to the third reservoir (R4) by making a round trip, - A ninth step (E9) of elution of the biological material by injecting the elution fluid from the first reservoir (RI) to the third reservoir (R4) by passing through the first space (100) of the filtration chamber (10), the filter (F) and the second space (101) of the filtration chamber (10).

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