Microfluidic system and method for processing a sample using the same

Abstract

The invention relates to a microfluidic cartridge (1) comprising a support in which a microfluidic circuit is made, 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 (R1) intended to receive 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).

Description

Technical field of the invention

[0001] The invention relates to a microfluidic system integrating a 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 out biological species present in the sample. To perform this filtration, the cartridge incorporates a filter designed 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 either fails to perform its function or requires the application of significant 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] The aim of the invention is therefore to propose a method for treating a fluidic sample, in which it is easy to unblock the filter, in case of the presence of bubbles on its surface. Description of the invention

[0005] This goal is achieved by a process for treating a fluidic sample comprising biological species, implemented in a microfluidic system, said microfluidic system comprising a microfluidic cartridge, actuation means and a control unit for said actuation means, the microfluidic cartridge comprising a support in which a microfluidic circuit is implemented, the microfluidic circuit comprising: A filtration chamber in which a filter is placed, said filter dividing 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; the microfluidic circuit comprising: 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 and second reservoirs each comprise a chamber and a deformable membrane within the chamber for moving a fluid. The actuation means are arranged and configured to control the movement of each membrane and the state of each valve in the microfluidic circuit. The process comprises: a first step of activating the opening of the first and second valves and the closing of the third valve; a second step of injecting the sample from the first reservoir into the second reservoir through the first opening of the filtration chamber; a third step of injecting the sample from the second reservoir into the first reservoir through the first opening of the filtration chamber; and a fourth step of activating 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 filtration chamber, so as to trap said biological species in the first space of the filtration chamber.

[0006] According to one particular feature, a cycle comprising the first command step, the second injection step and the third injection step is implemented several times, prior to the execution of the fourth command step.

[0007] According to another distinctive 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.

[0008] Another distinctive feature of the microfluidic circuit is that it includes: A third reservoir, 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,

[0009] The process also includes: A seventh control step at the closing of the third valve and the opening of the fourth valve, An eighth step of injection of an elution fluid from the third tank to the first tank or from the first tank to the third tank by making a round trip, A ninth step of elution of the biological material by injection of the elution fluid from the first tank to the third tank by passing through the first space of the filtration chamber, the filter and the second space of the filtration chamber.

[0010] Another objective of the invention is to propose a microfluidic system adapted to unblocking the filter and a method for implementing the unblocking of the filter.

[0011] This goal is achieved by a microfluidic system comprising a microfluidic cartridge, actuation means and a control unit for said actuation means, the microfluidic cartridge comprising a support in which a microfluidic circuit is implemented, the microfluidic circuit comprising: A filtration chamber in which a filter is placed, said filter dividing 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 and second reservoirs each comprise a chamber and a deformable membrane within the chamber to allow the movement of a fluid. The actuation means are arranged and configured to control the movement of each membrane and the state of each valve in the microfluidic circuit. The control unit is configured to command the actuation means according to a sequence, said sequence comprising several steps performed successively, each step being as defined in the treatment process above.

[0012] Depending on one particular feature, the microfluidic circuit includes: A third reservoir, 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, the sequence being executed by the control unit to implement the steps of the process defined above. Brief description of the figures

[0013] Other features and advantages will appear in the detailed description that follows, in conjunction with the attached drawings, in which: There figure 1 schematically illustrates the principle of implementation of the microfluidic system of the invention; The figure 2 shows the microfluidic cartridge of the invention in a top view; The Figures 3A and 3B illustrate the operating principle of a microfluidic capsule; The figures 4A to 4Cillustrate the different stages of the treatment process according to the invention; The figure 5 shows an example of the multilayer architecture of the microfluidic cartridge of the invention; Detailed description of at least one embodiment

[0014] With reference to the figure 1 and to the figure 2 The microfluidic system comprises: A microfluidic cartridge 1 in which a microfluidic circuit is made; Actuation means 2 for the microfluidic elements present in the microfluidic circuit, advantageously of the pneumatic type; A control unit UC intended to control the actuation means 2;

[0015] The microfluidic cartridge 1 is in the form of a support made of a transparent material, for example COC, PMMA or equivalent.

[0016] The support has a large surface area in two dimensions and a small thickness relative to those two dimensions.

[0017] The support structure is made of several layers sealed together. An architectural example will be described below in connection with the figure 5 The microfluidic circuit is integrated into the substrate by engraving, machining, embossing (or other known techniques) on one or more layers of the substrate. The same applies to the (pneumatic) control circuit used to control the fluidic elements of the cartridge.

[0018] The microfluidic circuit of a microfluidic cartridge includes several fluidic elements, including several reservoirs and several valves.

[0019] With reference to the figure 3A and to the figure 3BEach 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 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 movement of the membrane 30 within the chamber is controlled, and therefore 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 thus drawn into the first volume of the chamber (application of a negative pressure (P-) - figure 3A ) or push the fluid 31 out of the first volume of the chamber (application of a positive pressure (P+) - figure 3B ).

[0020] Each valve is also advantageously of the pneumatic type, controllable between an open state in which it allows fluid to pass through and a closed state in which it blocks the passage of fluid.

[0021] Within the framework of the invention, the microfluidic circuit comprises the following: A filtration chamber 10. This filtration chamber 10 includes a filter F dividing 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-restrictive manner and are to be understood when the cartridge is lying flat. A first reservoir R1 intended to receive 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 purge reservoir R2. 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.

[0022] The F filter is in the form of a grid, with a mesh whose pore size is adapted to the size of the biological species E to be trapped.

[0023] Within the framework of the invention, the first tank R1 and the second tank R2 have the architecture already described above.

[0024] In addition, the microfluidic circuit incorporates: A first valve V1 arranged on the first microfluidic channel C1 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;

[0025] 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.

[0026] This R4 tank has an identical architecture to that of the R1 and R2 tanks described above.

[0027] Each R1, R2 and R4 reservoir of the microfluidic circuit can 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.

[0028] As indicated above, the system incorporates 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.

[0029] It should be noted that the microfluidic cartridge can incorporate, within its support, a single layer forming a membrane common to all actuated fluidic elements (valves, reservoirs, etc.). By applying localized positive or negative pressure to this membrane, the corresponding fluidic element is actuated. This architecture is described in particular in connection with the figure 5 below.

[0030] The system also incorporates a control unit (CU) responsible for controlling the actuation means 2 to execute a sequence. The sequence comprises several steps performed 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.

[0031] Based on the system architecture described above, in conjunction with the figures 4A to 4C The processing method for the invention is as follows: The sample is initially placed in the first reservoir R1, this sample containing biological species E. E1 - Figure 4A The actuation means 2 are controlled to open the first valve V1 and the second valve V2, and to close the third valve V3. E2 - Figure 4AThe actuation means 2 are controlled to allow injection of the sample from the first reservoir R1 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. E3 - Figure 4A The actuation means 2 are controlled to allow the sample to be injected from the second reservoir R2 to the first reservoir R1 through the first opening 100 of the filtration chamber 10. The reverse process is then repeated, returning the sample to the first reservoir R1. This round trip cycle can be performed once or several times. Its purpose is to clean the filter F of any bubbles present on its surface and to unblock it, in preparation for the filtration of the sample. E4 - Figure 4BThe actuation means 2 are controlled to close the second valve V2 and open the third valve V3. The first valve V1 remains open. E5 - Figure 4B : The actuation means 2 are controlled for the injection of the sample from the first reservoir R1 to the third microfluidic channel C3, 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.

[0032] Before any filtration of a sample or any other compound, steps E1 to E3 can be advantageously implemented to unclog the filter and improve subsequent filtration. Thanks to the invention, complex analytical protocols using multiple samples and reagents can thus be performed. In particular, a washing liquid can be used, which is passed through the first space of the filtration chamber, between reservoirs R1 and R2, to wash and unclog filter F.

[0033] Furthermore, thanks to the system architecture of the invention, it is also possible to foresee the following additional steps: E6 - Figure 4B It is possible to perform 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). E7 - Figure 4CThe actuation means 2 are controlled to close the third valve V3 and open the fourth valve V2. The first valve remains open. E8 - Figure 4C The actuation means are controlled for injecting an elution fluid from reservoir R4 to the first reservoir R1, 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 C1. It would also be possible to inject the elution fluid from reservoir R1 to reservoir R4 and make several round trips until the elution fluid is recovered from reservoir R1. E9 - Figure 4CThe actuation means 2 are controlled for injecting the elution fluid from the first reservoir R1 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.

[0034] It should be noted that it would be possible to separate the volume recovered in tank R4 into several fragments, each fragment being collected in a separate tank.

[0035] There figure 5Figure 1 shows an example of a microfluidic cartridge. The microfluidic cartridge 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) forms the first space 100 of the filtration chamber 10. A second layer L2, forming the second space 101 of the filtration chamber, is then attached to this first layer L1, possibly with the aid of an adhesive layer (not shown) perforated in certain areas. This second layer L2 may also include several pneumatically actuated inlets. A third layer L3, forming the membrane, is then added to the second layer L2. This is a so-called full-plate membrane (judiciously perforated – particularly at 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, namely the reservoirs R1, R2, R4 and the valves V1, V2, V3, V4.

[0036] It should be noted that: Filter F is positioned between the first layer L1 and the second layer L2, separating the filtration chamber 10 into its two spaces 100 and 101, formed respectively in the first layer L1 and the second layer L2. Fluid passages are provided through the layers of the cartridge to connect the fluid elements of the fourth layer L4 to the two spaces 100 and 101 of the filtration chamber 10. The membrane is perforated at certain points, particularly at the filter, to allow the fluid to pass through filter F.

[0037] The invention thus offers numerous advantages, including: A simplified solution for unblocking a filter F from a microfluidic cartridge 1; A simple and reliable process for unblocking the filter F;

Claims

1. A method for processing a fluidic sample comprising biological species (E), implemented in a microfluidic system, said microfluidic system comprising a microfluidic cartridge, actuation means (2) and a control unit (CU) of said actuation means (2), The 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) 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 (R1) intended to receive 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),and a first valve (V1) arranged on the first microfluidic channel (C1), The microfluidic circuit comprising: - 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), - 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 (R1) and the second reservoir (R2) each comprising a chamber and a deformable membrane in the chamber to be able to move a fluid. - The actuation means (2) being arranged and configured to control the displacement of each membrane and the state of each valve of the microfluidic circuit, said process being, characterized in thatIt comprises: - A first step (E1) of control to open the first valve (V1) and the second valve (V2) and to close the third valve (V3), - A second step (E2) of injecting the sample from the first reservoir (R1) 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) to the first reservoir (R1) through the first space (100) of the filtration chamber, - A fourth step (E4) of control to close the second valve (V2) and open the third valve (V3), - A fifth step (E5) of injecting the sample from the first reservoir (R1) to 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,in order to trap said biological species (E) in the first space (100) of the filtration chamber.

2. Method according to claim 1, characterized in that a cycle comprising the first step (E1) of control, the second step (E2) of injection and the third step (E3) of injection is implemented several times, prior to the execution of the fourth step (E4) of control.

3. Method according to claim 1 or 2, characterized in that it includes a sixth step (E6) of lysis of biological species (E) trapped in the first space (10) of the filtration chamber, in order to release biological material (M).

4. Method according to claim 3, the microfluidic circuit (1) comprising: - A third reservoir (R4), - 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), Characterized in thatsaid process comprises: - A seventh step (E7) of control to the closing of the third valve (V3) and to the opening of the fourth valve (V4), - An eighth step (E8) of injection of an elution fluid from the third reservoir (R4) to the first reservoir (R1) or from the first reservoir (R1) to the third reservoir (R4) by making a round trip, - A ninth step (E9) of elution of the biological material by injection of the elution fluid from the first reservoir (R1) 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).

5. Microfluidic system comprising a microfluidic cartridge, actuation means (2) and a control unit (CU) of said actuation means (2), the microfluidic cartridge (1) having 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) 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 (R1) intended to receive 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), and a first valve (V1) arranged on the first microfluidic channel (C1), - 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), - 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 (R1) and the second reservoir (R2) each comprising a chamber and a deformable membrane within the chamber to allow the movement of a fluid. - The actuation means (2) being arranged and configured to control the movement of each membrane and the state of each valve in the microfluidic circuit, Characterized in that- The control unit is configured to control the actuation means according to a sequence, said sequence comprising several steps carried out successively, each step being as defined in the processing method of one of claims 1 to 3.

6. Microfluidic system according to claim 5, characterized in that The microfluidic circuit comprises: - A third reservoir (R4), - 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), and in that the sequence is executed by the control unit to implement the steps of the process defined in claim 4.

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