Metering microfluidic device and process for mixing two liquids in such a device

Abstract

This metering microfluidic device (2) comprises at least a first overflow reservoir (R1), a last overflow reservoir (R3), a meeting and merging cavity (32), a first control volume (Vc1) connected to the meeting and merging cavity, via a first primary connection (41), and to the first overflow reservoir, via a first secondary connection (51), and a last control volume (Vc3) connected to the meeting and merging cavity, via a last primary connection (43), and to the last overflow reservoir, via a last secondary connection (53). This metering microfluidic device also comprises a first injection port (i1) for injecting a first liquid to be mixed into the first control volume (Vc1), a last injection port (i3) for injecting a last liquid to be mixed into the last control volume (Vc3), a first entry port (e1) for injecting a first pushing gas into the first control volume (Vc1), and a last entry port (e3) for injecting a last pushing gas into the last control volume (Vc3).

Description

[0001] METERING MICROFLUIDIC DEVICE AND PROCESS FOR MIXING TWO LIQUIDS IN SUCH A DEVICE

[0002] TECHNICAL FIELD OF THE INVENTION

[0003] The present invention relates to a metering microfluidic device suitable for mixing and metering at least two liquids. The present invention also relates to a process for mixing at least two liquids in such a metering microfluidic device.

[0004] BACKGROUND OF THE INVENTION

[0005] Some applications require mixing of precise micro-quantities of different liquids. Such applications include industry in general and, more particularly, on-site testing, screening and / or diagnosis, as well as human or animal medical analysis laboratories. It happens sometimes that the initial volumes of some liquids to be mixed for such applications are not precisely known nor controlled. In such a case, it is complicated to realize the mixing of several liquids with given proportions.

[0006] For instance, in some cases, reagents have to be mixed with samples of body fluids. Consider the case where a concentration of a given compound in a body fluid, such as lithium in the saliva of a patient, must be determined. This can be used to assess whether the patient correctly follows a posology regarding a given medicament. In such a case, it is possible to collect a sample of saliva and to put it in contact with a given quantity of a reagent. However, the accuracy of the determination of the concentration of lithium in the example depends on the quantity of saliva put in contact with the reagent. If this quantity is neither known nor controlled, the accuracy of the determination might be unsatisfactory. This implies that a medical practitioner must perform the collection of saliva. This prevents the patient from performing a control by himself / herself.

[0007] Similar difficulties arise in other applications, as long as the volume of at least one micro-quantity of liquid is not precisely known nor controlled. Such other applications include the control of the quality of potable water, the quality control of products in food industry, chemical industry or biological industry.

[0008] In the article “A capillary-pressure-based air pump for nanofilter liquid handling in microfluidic devices” (15thInternational Conference on Miniaturized Systems for Chemistry and Life Science, October 2-6, 2011 , Seattle, Washington, USA), Li et al disclose a microfluidic pump for mixing two liquids. This pump is limited to two liquids and mixing is based on the assumption that the volume of the two liquids is known in advance. On the other hand, in the article “Microfluidic chain reaction of structurally programmed capillary flow events” (Nature - vol 605 - page 464 - 19 May 2022), Yafia et al describe a mixing device equipped with valves whose operation depends on the knowledge of the fluids to be merged. In the article “Integration en consommable plastique jetable: Fabrication, essais fluidiques et performance biologique” ( La Houille Blanche I N° 4-2003), Broyer et al consider different types of microfluidic devices for us with chemical compounds whose quantity is a priori known in advance. In the three above-mentioned documents, one relies on a capillarity effect for the dosing of the products within the fluidic devices, which limits the possibilities of use of these devices.

[0009] In the article “3D-printed capillary ELISA-on-a-chip-lp with aliquoting” (bioRxiv preprint doi September 26, 2022), Parandakh eta / disclose chip based devices, in particular devices including reagents capable of reacting with a saliva sample, where different liquids are sequentially injected within the chip, in a known quantity.

[0010] EP3779439B1 discloses a microfluidic chip and an analysis instrument including such a chip where a liquid quantification chamber has a predetermined volume, whereas a liquid to be quantitatively determined flows into the liquid quantification chamber. Injection of the liquid to be quantitatively determined is tricky since it is made by capillarity. EP3505249B1 discloses a sample loading system and a method for metering a predetermined amount of sample, based on a precise measuring of the volume of the fluid sample. This is not always possible, in particular for samples of body fluids or samples taken in an industrial environment. US2015 / 0147777A1 discloses a passive microfluidic metering device that meters successive volumes of a fluid for performing a series of metering operations. This device is complicated to operate and does not allow mixing different fluids of unknown volumes. US2023 / 0285964A1 discloses several metered volume microfluidic devices, which comprise a mixing chamber including a reactant. These devices allow metering a single volume of fluid, the volumes of the other fluid interacting with the metered fluid need to be known in advance. EP1942347A1 discloses a cartridge for analyzing blood, which includes where blood and air are introduced via a same hole. Air is also injected in another hole. On the other hand, US2007 / 0111302A1 discloses a device for controlling the size and movements of some microdroplets, which does not make use of reservoirs or control volumes.

[0011] SUMARRY OF THE INVENTION

[0012] Thus, there is a need for a metering microfluidic device capable of handling different liquids whose volumes are not precisely known nor controlled, in a simple, cost effective and reliable manner.

[0013] To this end, the present invention concerns a metering microfluidic device comprising at least: - a first overflow reservoir and a last overflow reservoir;

[0014] - a meeting and merging cavity;

[0015] - a first control volume connected to the meeting and merging cavity, via a first primary connection, and to the first overflow reservoir, via a first secondary connection;

[0016] - a last control volume connected to the meeting and merging cavity, via a last primary connection, and to the last overflow reservoir, via a last secondary connection;

[0017] - a first injection port for injecting a first liquid to be mixed into the first control volume;

[0018] - a last injection port for injecting a last liquid to be mixed into the last control volume;

[0019] - a first entry port for injecting a first pushing gas into the first control volume; and

[0020] - a last entry port for injecting a last pushing gas into the last control volume.

[0021] Thanks to the invention, the first and last overflow reservoirs allow accommodating quantities of liquids to be metered, in excess of the first and second control volumes. This allows mixing the liquids in proportions determined by the ratio of the capacities of the respective control volumes, without precisely knowing the quantities of the samples of liquid to be mixed and metered. On the other hand, the injection points and the entry points allow controlling the flow of liquids to be mixed and metered into the control volumes and, from there, into the overflow reservoirs and into the meeting and merging volume. In addition, the device of the invention may have no valve to control the flow of liquids to be mixed, which simplifies this device.

[0022] According to advantageous and non-compulsory aspects of the invention, such a metering microfluidic device may incorporate one or several of the following features taken in any technically admissible configuration:

[0023] - The metering microfluidic device includes a curved surface forming a smooth vertical transition zone between each secondary connection and the adjacent overflow reservoir, with a shape such that no air gap is formed above a quantity of liquid present in an overflow reservoir.

[0024] - Each overflow reservoir is equipped with a vent port.

[0025] - The capacity of each overflow reservoir is larger than the capacity of the control volume connected to this reservoir.

[0026] - The metering microfluidic device includes a homogenization area located downstream of the meeting and merging cavity and a mixture collection zone located downstream of the homogenization area.

[0027] - A fluidic resistance of a secondary connection is smaller than a fluidic resistance of the primary connection connected to the same control volume. - An outlet surface of a secondary connection into the corresponding overflow reservoir has no discontinuities and is preferably represented by a continuously differentiable function.

[0028] - An outlet of a primary connection into to the meeting and merging cavity is configured to hold a flow of liquid to be mixed within the control volume connected to this primary connection.

[0029] - The meeting and merging cavity is defined between at least two opposite walls, an outlet of any primary connection but the last one is located on a first wall among the two opposite walls and configured so that the liquid to be mixed coming out of the primary connection adheres on the first wall without contacting the second wall and the outlet of the last primary connection is configured so that the last liquid to be mixed contacts the two opposite walls when coming out of the last primary connection.

[0030] - A downstream outlet of any primary connection but the last one is of the curvecurve type, of the right-curve type or of overhanging curve-curve type, whereas a downstream outlet of the last primary connection is defined by a sharp edge and centered between the two opposite walls of the meeting and merging cavity.

[0031] - The microfluidic device has no valve for controlling the flow of liquids to be mixed, into and out of the control volumes.

[0032] - The metering microfluidic device is formed by a stack of plates which define parts of the overflow reservoirs, parts of the control volumes and / or parts of the injection or entry ports.

[0033] - The metering microfluidic device is monobloc, preferably made by 3D printing, stereo photolithography, injection molding or sintering.

[0034] According to a second aspect, the invention relates to a process for mixing at least two liquids in a metering microfluidic device as previously described, comprising at least the following steps: a) injecting the first liquid to be mixed into the first control volume, via the first injection port, until a portion of the first liquid enters the first overflow reservoir via the first secondary connection; b) injecting the last liquid to be merged into the last control volume, via the last injection port, until a portion of the last liquid enters the last overflow reservoir via the last secondary connection; c) injecting the first pushing gas into the first control volume, via the first entry port, until all the first liquid to be mixed contained in the first control volume is pushed by the first pushing gas into the meeting and merging cavity, via the first primary connection; and d) injecting the last pushing gas into the last control volume, via the last entry port, until all the last liquid to be mixed contained in the last control volume is pushed by the last pushing gas into the meeting and merging cavity, via the last primary connection.

[0035] This process induces the same advantages as the device of the invention.

[0036] Advantageously, the pushing gas is the same for all control volumes.

[0037] BRIEF DESCRIPTION OF THE DRAWINGS

[0038] The invention will be better understood, based on the following description of a metering microfluidic device and an associated mixing process according to the invention, given as an illustrative example only, in reference to the annexed drawings where:

[0039] - Figure 1 is a schematic front view of a microfluidic device according to the invention;

[0040] - Figure 2 represents, on three inserts A), B), C), an exploded view, a partial cut view and an enlarged view of detail C on figure 1 ;

[0041] - Figure 3 represents several steps of a process according to the invention, for mixing three liquids; and

[0042] - Figure 4 represents on two inserts A and B some control volumes of the device of figures 1 and 2 and some possible geometries of a downstream outlet of a connection used in the device of figures 1 and 2.

[0043] DETAILED DESCRIPTION OF SOME EMBODIMENTS

[0044] A single-use metering microfluidic device 2 according to the invention is represented on figure 1 and configured for mixing three liquids, in particular liquids. Hereafter, the metering microfluidic device 2 may also be identified as “the microfluidic device 2” or as “the device 2”.

[0045] The metering microfluidic device 2 is made of three plates 22, 24 and 26, stacked together with the plate 24 located above the plate 22 and the plate 26 located above the plate 24. A bottom liner 28 is attached to a bottom surface of the first plate 22 opposite to the second plate 24. The three plates 22, 24 and 26 and the liner 28 together form a stack 30, which constitutes the metering microfluidic device 2 and defines several volumes, connections, injection points, entry points and vents, as explained here below.

[0046] Preferably, the plates 22, 24 and 26 are at least partly transparent.

[0047] Each plate 22, 24 and 26 can be made by injection molding of a synthetic material, laser cutting or embossing of a metallic sheet. Other techniques are also possible.

[0048] The bottom liner is made by a sheet of either single face sticky tape or contact tape. For the sake of simplicity, the bottom liner 28 is represented only on insert B) of figure 2. According to non-represented alternative embodiments of the invention, the metering microfluidic device 2 can be monobloc, for instance made by 3D printing, stereophotolithography, injection molding or sintering.

[0049] Injection molding, sintering and / or embossing can be used for mass production of large quantities of metering microfluidic devices 2. Laser cutting and / or 3D printing can be used for small-series or on-demand production, for instance for academic research.

[0050] Several volumes, connections and entry / exit ports for fluids are defined by the three plates 22, 24 and 26 and the bottom liner 28. All the volumes, connections and ports mentioned here below are formed by through-holes made in one or several of the plates 22, 24 and 26.

[0051] The plate 22 defines a first portion R12of a first overflow reservoir R1. The first plate 22 also defines a first portion R22of a second overflow reservoir R2 and a first portion R32of a third overflow reservoir R3.

[0052] The second plate 24 defines second portions R14, R24and R34of the overflow reservoirs R1 , R2 and R3.

[0053] As shown for the first overflow reservoir R1 on insert B) of figure 2, the first reservoir R1 is formed by its portions R12and R14located between the third plate 26 and the bottom liner 28.

[0054] The device 2 also defines three control volumes Vc1 , Vc2 and Vc3.

[0055] These three volumes Vc1 , Vc2 and Vc3 are formed by through-holes made in the first plate 22 and extend between the second plate 24 and the bottom liner 28.

[0056] The device 2 also defines a meeting and merging cavity 32, a homogenization area 34 and a mixture collection zone 36, all made by through holes in the first plate 22 and located between the second plate 24 and the liner 28.

[0057] The meeting and merging cavity 32 is connected to the three control volumes Vc1 , Vc2 and Vc3, respectively by a first primary connection 41 , a second primary connection 42 and a third primary connection 43. These connections are formed by through-holes provided in the first plate 22.

[0058] When liquids are present in the meeting and merging cavity 32, they can be pushed through the homogenization area 34 up to the mixture collection zone 36. Thus, in the direction of mixed liquids, the homogenization area 34 is located downstream of the meeting and merging cavity 32 and the mixture collection zone 36 is located downstream of the homogenization area 34.

[0059] Secondary connections are also provided between the control volumes and the overflow reservoirs. More precisely, a first secondary connection 51 connects the first control volume Vc1 to the first overflow reservoir R1 , a second secondary connection 52 connects the second control volume Vc2 to the second overflow reservoir R2 and a third secondary connection 53 connects the third control volume Vc3 to the third overflow reservoir R3.

[0060] In the example of the figures, three control volumes, three overflow reservoirs, three primary connections and three secondary connections are provided, so that the third control volume Vc3, overflow reservoir R3 primary and secondary connections 43 and 53 are respectively a last control volume, a last overflow reservoir and last primary and secondary connections.

[0061] In the example of the figures, the three liquids to be mixed are liquids.

[0062] In the example of the figures, the number N of control volumes Vc1 , Vc2 and Vc3 equals three.

[0063] In alternative embodiments of the invention, this number N can be equal to two or larger than or equal to four. Advantageously, the number of overflow reservoirs, primary connections and secondary connections is the same as the number of control volumes.

[0064] The control volume, the overflow reservoir, the primary connection and the secondary connection with rank N form the last control volume, the last overflow reservoir, the last primary connection and the last secondary connection. Depending on the value of N, zero, one or more control volumes, overflow reservoirs, primary and secondary connections may be interposed between the first and last control volumes, overflow reservoirs, primary and secondary connections.

[0065] Generally speaking, the number N equals the number of liquids to be mixed in the metering microfluidic device 2.

[0066] As shown by the curved surface S24T visible on insert B) of figure 2, an edge of the second plate 24 adjacent to the second portion R14of the first overflow reservoir R1 is rounded, so that it constitutes a smooth vertical transition zone between the first secondary connection 51 and the first overflow reservoir R1 . The curved surface S24T is smooth, in so far as it has no discontinuity. Preferably, this surface is represented by a continuously differentiable function. Preferably, the curved surface S24T has a capillary effect to effectively guide a liquid into the second portion R14of the first overflow reservoir R1. Here, the capillary effect can be defined as favourable conditions for the liquid to move toward the second portion R14with very low pressure. Texturing of the curved surface S24T increases the liquid's contact surface with the metering microfluidic device 2, and thus favours liquid movement, because wettability is increased, like with blotting paper. In this way, the mass of liquid in contact with the channel-reservoir junction is sufficient to generate a fluidic resistance preventing the formation of an air gap above the liquid. This allows effectively controlling the quantity of the first liquid sent to the meeting and merging cavity 32, as explained below.

[0067] Similar transition zones also exist for the second and third overflow reservoirs R2 and R3. All these transition zones are represented by dark areas on the right of the second portions R12, RI4 and R16in the central zone of insert A) of figure 2.

[0068] According to an advantageous aspect of the invention, the shape of each curved surface S24T or equivalent is such that the liquid in each overflow reservoir R1 , R2 or R3 is in contact with both the liner 28 and the third plate 26, beyond the surface S24!, as shown on insert B) of figure 2. This implies that no air gap exists above the liquid present in each overflow reservoir.

[0069] The metering microfluidic device 2 includes a first injection port i1 for injecting into the first control volume Vc1 a first liquid to be mixed. This injection port i1 is located near the first primary connection 41 and extends through the three plates 22, 24 and 26. More precisely, the first injection port i1 is formed of three through-holes i12, i14and i16respectively made into the three plates 22, 24 and 26.

[0070] The metering microfluidic device 2 also includes a second injection port i2 and a third and last injection port i3, respectively located near the second and third primary connections 42 and 43. Each injection port i2 and i3 is also made of three through-holes i22, i24and i26and i32, i34and i36respectively provided in the three plates 22, 24 and 26.

[0071] The metering microfluidic device 2 also includes a first entry port e1 for injecting a first pushing gas into the first control volume Vc1 . The first entry port e1 is located near the secondary connection 51 .

[0072] Similarly, a second entry port e2 and a third and last entry port e3 are provided for injecting a pushing gas into the second and third control volumes Vc2 and Vc3. These second and third entry ports are respectively located near the second and third secondary connections 52 and 53.

[0073] Advantageously, the entry port e1 is made by three through-holes e12, e14and e16, respectively provided in the plates 22, 24 and 26. Similarly, the entry ports e2 and e3 are made of three through-holes e22, e24and e26and e32, e34and e36respectively provided in the plates 22, 24 and 26.

[0074] Advantageously, the pushing gas is the same for all three control volumes, preferably a gas. In the example of the figures, this gas is air.

[0075] The metering microfluidic device 2 also include three vent ports v1 , v2 and v3. Vent port v1 is formed by a through-hole in the third plate 26 and connects the first overflow reservoir R1 to the atmosphere. Similarly, vent ports v2 and v3 are made by through-holes in the third plate 26 and connect the second and third overflow reservoirs R2 and R3 to the atmosphere. The metering microfluidic device 2 also includes a terminal vent port v36 made by through-holes v634and v366provided in the second and third plates 24 and 26 and which connects the collection zone 36 to the atmosphere.

[0076] The vent, injection and entry ports are sealed by stoppers, as shown with stopper 61 in insert B) of figure 2. They are opened by removing the stoppers, when necessary, to allow fluid motion, as explained here below.

[0077] As an example, the three plates can be of a general square shape of 65 mm by 65 mm. The third plate 26 is advantageously thicker than the first and second plates 22 and 24 since the through-holes made in this plate are designed to accommodate a tip of a pipette, as explained here above, or a stopper.

[0078] The first plate 22 contains the general fluidic circuit of the device 2. The height or thickness of this circuit is about 1 .6 mm and the lateral dimensions of the channels provided in this plate are 0.5 mm for channels 41 , 42, 43 and 1 mm for channels 51 , 52, 53 in this particular example. Plates 22 and 24 may have the same thickness, i.e. 1.6 mm, whereas the third plate 26 is thicker. For instance, the overall thickness T30of the stack 30 may be equal to 7.5 mm.

[0079] Operation of the metering microfluidic device 2 is based on the controlled management of the fluidic resistances existing between this device and the liquids to be mixed therein. These fluidic resistances depend on the cross-sections of the fluid channels used in each step of a process for moving and mixing the three liquids within this device, as explained here below. These fluidic resistances also depend on the geometry of some outlets and on some contact angles of the liquids with the surface of these fluid channels.

[0080] This process is also for metering the three liquids L1 , L2 and L3.

[0081] The capacities of the controlled volumes Vc1 , Vc2 and Vc3 are respectively equal to 30 pL, 40 pL and 50 pL.

[0082] In a variant, the capacities may be different. Advantageously, in such a case, the capacity of the third and last control volume Vc3 is the largest one.

[0083] Figure 3 illustrates the operation of the mixing microfluidic device 2 for mixing of three liquids, for instance three liquids.

[0084] On figure 3, when one port is black, it means that this port is closed, either by a stopper or by a pipette. When a port is white, it means that this port is open, that is non-obstructed.

[0085] At the beginning of the process for mixing three liquids represented on figure 3, all ports are closed, as shown by the first step represented on the top left corner of figure 3.

[0086] Then, as shown in the second step represented at the center of the first row of figure 3, port i1 and v1 are opened and a first liquid L1 is injected via the injection port i1 into the control volume Vc1 . The quantity of this first liquid does not have to be precisely known, in so far as it is above 30 pL, since the first liquid in excess flows from the control volume Vc1 into overflow reservoir R1. The flow of the first liquid toward the overflow reservoir R1 is facilitated by the fact that the first vent v1 is opened, whereas the vent v36 is closed. This is also facilitated by the geometry of the first primary and secondary connections 41 and 51. The flow of the first liquid L1 toward the overflow reservoir R1 can be monitored by looking at the overflow reservoir through the third plate 26, which is partly transparent, or by controlling the duration of injection of the first liquid L1.

[0087] Thereafter, second and third liquids L2 and L3 are successively and respectively injected into the second and third control volumes Vc2 and Vc3, via the injection ports i2 and i3, whereas the vent port v2 and v3 are open. In those cases, the liquids L2 and L3 in excess flow into the overflow reservoirs R2 and R3, as shown in the third step represented on the top right corner of figure 3. The flow of the second and third liquids L2 and L3 toward the overflow reservoirs R2 and R3 can be monitored by looking at the overflow reservoirs through the third plate 26, which is partly transparent, or by controlling the duration of injection of the second and third liquids L2 and L3.

[0088] According to a non-represented variant of the invention, injection of the liquids L1 , L2 and L3 in the control volumes Vc1 , Vc2 and Vc3 can occur in an order different from the one mentioned here above, via the three injection ports i1 , i2 and i3.

[0089] Then, as shown in the fourth step represented on the left of the center row of figure 3, all ports are closed apart from the entry port e1 and the vent port v36.

[0090] The pushing gas may be injected into the control volume Vc1 through a micropipette 100, such as the one partly represented on insert B) of figure 2, inserted into the entry port e1. The flow of pushing gas is shown by arrow A1 on insert B) of figure 2. Alternatively, the pushing gas is inserted in the control volume Vc1 by means different from a pipette, such as an external injection pump equipped with valves, which forms a source of pushing gas.

[0091] Pushing gas injection via the first entry point e1 takes place until all the first liquid L1 contained in the first control volume Vc1 is pushed by the pushing gas into the meeting and merging cavity 32, via the first primary connection 41. This can be monitored by looking at the first control volume Vc1 through the second and third plates 24 and 26, which are transparent.

[0092] Since the vent port v1 is closed whereas the vent port v36 is opened and due to the geometry of the first primary and secondary connections 41 and 51 , the first liquid L1 originally contained into the control volume Vc1 is pushed, by the pushing gas coming out of the pipette 100 or from the external injection pump, into the meeting and merging cavity 32 where it adheres on a first wall 322 of this cavity where an outlet of the first primary connection 41 is located. This allows pouring a quantity Q1 of the first liquid L1 into the meeting and merging cavity 32. This first quantity Q1 is defined by the capacity of the first control volume Vc1.

[0093] The same operation is implemented for the second liquid L2, as shown by the fifth step represented in the center of the second row of figure 3. The pushing gas is injected into the second control volume via the second entry port e2. Here, a second quantity Q2 of the second liquid L2 is ejected from the second control volume Vc2 into the meeting and merging cavity 32 and adheres to the first wall 322. This second quantity Q2 is defined by the capacity of the second control volume Vc2.

[0094] The same operation is also implemented for the third liquid L3, as shown in the sixth step represented on the right of the central row of figure 3. The pushing gas is injected into the third control volume via the third entry port e3. Here, a third quantity Q3 of the third liquid L3 is ejected from the third control volume Vc3 into the meeting and merging cavity 32 and adheres to the first wall 322 and to an opposite wall 324 of the meeting and merging cavity 32. This third quantity Q3 is defined by the capacity of the third control volume Vc3.

[0095] In the microfluidic device 2, for j an integer between 1 and 3, each entry port ej used for injecting the pushing gas into the control volume Vcj, is different from the injection port ij used for injecting the liquide to be mixed into the same control volume Vcj. Actually, the injection port ij and the entry port ej associated with a control volume Vcj are located at two opposite ends of this control volume.

[0096] In the fifth and sixth steps, pushing gas injection via the second and third entry points e2, e3 takes place until all the second liquid L2 contained in the second control volume Vc2 and all the third liquid L3 contained in the third control volume Vc3 is pushed by the pushing gas into the meeting and merging cavity 32, via the second or third primary connection 42, 43. This can be monitored by looking at the second and third control volumes Vc2 and Vc3 through the second and third plates 24 and 26, which are transparent, or by controlling the duration of injection of pushing gas.

[0097] In the fourth, fifth and sixth steps, the pushing gas does not flow into the overflow reservoirs R1 , R2 and R3 since no air gap exist above the liquid present in these reservoirs, due to the shape of the curved surface S24!, for the first overflow reservoir R1 , and of equivalent surfaces for the other two overflow reservoirs R1 and R2. Thus, the pushing gas is efficient to push the liquid L1 , L2 or L3 out of the corresponding control volume Vc1 , Vc2 or Vc3.

[0098] The fourth, fifth and sixth steps can occur in any order as long as the liquid quantity Q3 does not enter in contact with the primary connection 42.

[0099] In a seventh step represented on the lower left corner of figure 3, the pushing gas injected via the third entry port e3 into the third control volume Vc3 flows into the meeting and merging cavity 32 and pushes the three quantities Q1, Q2 and Q3 of liquids L1 , L2 and L3 toward a downstream end of the meeting and merging cavity 32, which merges these liquid quantities Q1, Q2 and Q3 and moves them into the homogenization area 34.

[0100] In an eighth step represented at the center of the lower row of figure 3, the blend of the three liquids circulates through the homogenization area 34, which results in an intimate mixing of the three liquid quantities Q1, Q2 and Q3. The blend of liquids is pushed by the pushing gas injected into the third entry port e3, which goes all the way through the meeting and merging cavity 32.

[0101] In a ninth step of the method represented in the bottom right corner of figure 3, the mixture M of liquids reaches the mixture collection zone 36 from where it can be collected via the terminal vent v36, while being pushed by the pushing gas injected into the third entry port e3. Alternatively, the mixture M may remain in the mixture collection zone 36 if any sensing / analysis / other operation is performed here on this mixture.

[0102] Thus, thanks to the use of the control volumes Vc1 , Vc2 and Vc3 and of the injection and entry ports i1 , i2, i3, e1 , e2 and e3, it is possible to mix quantities Q1, Q2 and Q3 of liquid L1 , L2 and L3 which are equal to the respective capacities of the control volumes Vc1 , Vc2 and Vc3, irrespective of the actual quantity of liquids introduced in the metering microfluidic device 2 at the beginning of the mixing process of the invention, provided that the quantities of liquids introduced are larger than the respective capacities of the control volumes Vc1 , Vc2 and Vc3.

[0103] Insert A) of figure 4 shows, on the one hand, the geometry of the first two control volumes Vc1 and Vc2 and, on the other hand, the geometry of the third and last control volumes Vc3.

[0104] The two control volumes Vc1 and Vc2 have globally the same shape, even if they have different capacities, respectively 30 pL and 40 pL in this particular case.

[0105] The first and second control volumes Vc1 and Vc2 include a reservoir 90, a first tube 92 joining the reservoir to the first or second entry point e1 or e2 and a second tube 94 joining the reservoir to the first or second primary connection 41 or 42.

[0106] As can be deduced by the comparison of insert C) of figure 2 and insert A) of figure 4, the first liquid occupies more than the first control volume Vc1 after the second step of the process represented on figure 3. More precisely, after this step, the first liquid also occupies a tube 96 extending between the first injection port i1 and the second tube 94. The first liquid also occupies the volume of the first secondary connection 51 and a portion of the first overflow reservoir R1 . These volumes do not belong to the first control volume Vc1 and they are not emptied by the injection of pushing gas through the entry port e1 in the fourth step mentioned here above. As visible on the right of insert C) of figure 2, a sharp transition is provided between the downstream end of the second tube 94 and the upstream end of the first primary connection 41. This sharp transition is made by an annular surface Si, which is perpendicular to a longitudinal direction of the second tube 94 and which surrounds its downstream end. Due to the surface tension of the liquid present in the control volume Vc1 at this stage, this sharp transition induces a relatively high fluidic resistance, which holds the first liquid within the control volume Vc1 and resists flowing of the first liquid L1 toward the first primary connection 41 and toward the meeting and merging cavity 32.

[0107] On the other hand, the transition between the first secondary connection 51 and the first overflow reservoir R1 is formed by a smooth surface S2, which extends around a downstream end of the first secondary connection 51. This surface S2is smooth, in so far as it has no discontinuity. Preferably, this surface is represented by a continuously differentiable function. This second surface S2induces less fluidic resistance to the flow of the first liquid L1 toward the first overflow reservoir R1 than the sharp transition represented by the first surface ST and its sharp edge joining the second tube 94. Thus, the fluidic resistances of the first and second primary connections 41 and 42 are respectively higher than the fluidic resistances of the first and second secondary connections 51 and 52. This contributes to the fact that the liquid injected in the second step of the method tends to flow toward the first overflow reservoir R1 instead of toward the meeting and merging cavity 32.

[0108] The third control volume Vc3 includes a reservoir 90', a first tube 92' connecting it to the third entry port e3 and a second tube 94' connecting it to the third primary connection 43 and to the meeting and merging cavity 32. In the example of the figures, the third primary connection 43 is the downstream outlet of the second tube.

[0109] An annular surface S3surrounds the downstream end of the second tube 94', in the same way as the first surface ST for the first two control volumes Vc1 and Vc2. The shape of the transition between the third control volume Vc3 and the third overflow reservoir R3 is as explained here above for the first two control volumes. This also induces that, due to the surface tensions of the third liquid L3, the fluidic resistance to the flow toward the meeting and merging cavity 32 is higher than the fluidic resistance to the flow toward the third overflow reservoir R3. Thus, the fluidic resistance of the third primary connection 43 is higher than the fluidic resistance of the third secondary connection 53.

[0110] Thanks to the handling of the flows of liquids L1 , L2 and L3 via the respective fluidic resistances of the primary and secondary connections, it is not necessary to implement valves in the metering microfluidic device 2. In other words, the metering microfluidic device 2 has no valve for controlling the flow of liquids to be mixed, into and out of the control volumes Vc1 , Vc2 and Vc3. In addition, the metering microfluidic device 2 has no valve for controlling the flow of pushing gas, into and out of the control volumes Vc1 , Vc2 and Vc3. The role of valves is performed by the pipettes or by the valves of the external injection pump. Thus, the metering microfluidic device 2 does not need internal valves, which is very advantageous in terms of simplicity and cost for such a single-use device;

[0111] As shown on figure 3, the first two liquids reach the meeting and merging cavity 32 on its first wall 322, where they adhere, without contacting the second wall 324. This avoids entrapping the pushing gas or another fluid between the three quantities Q1, Q2 and Q3 of liquids arriving in the meeting and merging cavity 32.

[0112] This is obtained by the geometry of the downstream end of the first and second primary connections 41 and 42 which are shaped in order to make the quantities Q1 and Q2 adhere onto the first wall 322.

[0113] With the geometry represented with letter (a) on insert B) of figure 4, and which is of the “right-right” type, it cannot be guaranteed that the liquid flowing out of the first primary connection 41 will adhere to the first wall 322 without liquid in the primary connection 41 anymore. The situation is better for the junction of letter (b), which is of the “curve-curve” type where the downstream end of the first primary connection 41 is surrounded by two curved portions. The situation is also better with the junction represented with letter (c) which is of the “right-curve” type. The situation is optimal with the geometry represented with letter (d), which is of the “overhanging curve-curve” type, where the flow of liquid coming out of the first primary connection is efficiently directed toward the first wall 322 of the cavity 32. Advantageously, the quantity Q1 adheres to the wall 322 in the downstream direction of the cavity 32.

[0114] On the contrary, the geometry of the third primary connection 43 is defined by a sharp edge, between the downstream end of the second tube 94’ and the annular surface S3, as mentioned here-above for what concerns the fluidic resistance due to the surface tension. Moreover, this downstream end is centered between the two walls 322 and 324 of the cavity 32, so that the quantity Q3 of the third liquid L3 coming from the third control volume Vc3 flows in contact with these two walls, into the cavity 32.

[0115] This allows pushing the three quantities Q1 , Q2 and Q3 into the homogenization area 34 with pushing gas coming from the last entry ports e3.

[0116] Since the capacity of the third and last control volume Vc3 is advantageously the largest one between the capacities of all control volumes, the quantity Q3 is the largest one, which facilitates the fact that the quantity Q3 coming out of the third and last control volume Vc3 will adhere to the two opposite walls 322 and 324 of the meeting and merging cavity 32. According to an advantageous aspect of the invention, the walls 322 and 324 are smooth and are provided with a continuous curvature which facilitates adherence of the liquid, respectively on the first wall 322 for quantities Q1 and Q2 or on both walls 322 and 324 for quantity Q3.

[0117] The shape of the surface S24T allows the first liquid to adhere to this surface in the configuration represented on insert B) of figure 2, so that no pushing gas coming from the pipette 100 or from the external injection pump can penetrate into the first overflow reservoir R1 , which could be the case if no liquid were present in the second portion R14of the first overflow reservoir R1. In other words, the geometry of the surface S244, which is smooth and does not present discontinuities with regards to the surface of the second plate 24 defining the first secondary connection 51 , allows liquid to be present on the whole height of the first overflow reservoir R1 , in the zone represented on inserts B) and C) of figure 2. This avoids leakage of pushing gas into this reservoir and guarantees that the flow of pushing gas A1 is efficiently used to push the first liquid L1 out of the first control volume Vc1 , as explained here-above.

[0118] The same situation arises for the second and third control volumes and their transitions with the second and third overflow reservoirs.

[0119] Advantageously, generally speaking, the capacity of each overflow reservoir is larger than the capacity of the control volume connected to this reservoir. Actually, the capacity of each overflow reservoir is selected depending on the expected volume of the samples to be mixed and metered. This allows injecting relatively large quantities of liquid to be mixed and metered within the device 2, without specific precautions, whereas only small quantities Q1 , Q2 and Q3, namely 30 pL, 40 pL and 50 pL in this example, are used for the actual merging and mixing, that is the quantities defined by the control volumes.

[0120] In theory, there is no upper limit for the capacities of the overflow reservoirs R1 , R2 and R3. Regarding their lower limits, an important criterion is that an overflow reservoir must not be fully filled with liquid since, in such a case, the vent v1 , v2 or v3 could be clogged by the liquid, which could force the liquid to flow into the meeting and merging cavity 32, without control.

[0121] In addition, for i an integer between 1 and 3, in order to guarantee that only the quantity of liquid present in a control volume Vci flows into the meeting and merging cavity 32, the mass of liquid present between the entry point ei and the adjacent overflow reservoir Ri must be larger than or equal to the mass of the liquid present in the control volume Vci.

[0122] The homogenization area 34 is represented on the figures as a serpentine. However, other geometries and other types of homogenization can be contemplated, for instance, by application of the principles of the articles “Microfluidic Mixing: A Physics Oriented Review” (Micromachines 2023, 14, 1827) by Saravanakumar et al, “Mixing in Turbulent Flows: An overview of Physics and Modelling” (Processes 2020, 8, 1379) by Pozorski et al and “Microfluidic Mixing: A review” (International Journal of Molecular Sciences 2011 vol 12, p 3265) by Lee et al. The invention is described here above in case the metering microfluidic device 2 is designed for mixing and metering three liquids. In non-represented variants, this device can be designed for mixing two liquids only, or a number of liquids larger than or equal to four. The same applies for the process of the invention. The number N of control volumes, overflow reservoirs, injection ports and entry port is adapted to the number of liquids to be mixed.

[0123] In a non-represented variant of the invention, the pushing gas can be different from air. For instance, this pushing gas may be an inert gas, such as nitrogen, or a rare gas.

[0124] The embodiment, variants and optional aspects listed here-above may be combined to generate other embodiments of the invention defined in the appended claims.

Claims

CLAIMS1. A metering microfluidic device (2) characterized in that it comprises at least: a first overflow reservoir (R1) and a last overflow reservoir (R3); a meeting and merging cavity (32); a first control volume (Vc1) connected to the meeting and merging cavity (32), via a first primary connection (41), and to the first overflow reservoir (R1), via a first secondary connection (51); a last control volume (Vc3) connected to the meeting and merging cavity (32), via a last primary connection (43), and to the last overflow reservoir (R3), via a last secondary connection (53); a first injection port (i1) for injecting a first liquid to be mixed into the first control volume (Vc1); a last injection port (i3) for injecting a last liquid to be mixed into the last control volume (Vc3); a first entry port (e1) for injecting a first pushing gas into the first control volume (Vc1); and a last entry port (e3) for injecting a last pushing gas into the last control volume (Vc3).

2. The metering microfluidic device of claim 1 , including a curved surface (824^ forming a smooth vertical transition zone between each secondary connection (51 , 52, 53) and the adjacent overflow reservoir (R1 , R2, R3), with a shape such that no air gap is formed above a quantity of liquid present in an overflow reservoir..

3. The metering microfluidic device of any preceding claim, wherein each overflow reservoir (R1 , R2, R3) is equipped with a vent port (v1 , v2, v3).

4. The metering microfluidic device of any preceding claim, wherein the capacity of each overflow reservoir (R1 , R2, R3) is larger than the capacity of the control volume (Vc1 , Vc2, Vc3) connected to this reservoir.

5. The metering microfluidic device of any preceding claim, wherein it includes a homogenization area (34) located downstream of the meeting and merging cavity (32) and a mixture collection zone (36) located downstream of the homogenization area.

6. The metering microfluidic device of any preceding claim, wherein a fluidic resistance of a secondary connection (51 , 52, 53) is smaller than a fluidic resistance of the primary (41 , 42, 43) connection connected to the same control volume (Vc1 , Vc2, Vc3).

7. The metering microfluidic device of claim 6, wherein an outlet surface (S2) of a secondary connection (51 , 52, 53) into the corresponding overflow reservoir (R1 , R2, R3)has no discontinuities and is preferably represented by a continuously differentiable function.

8. The metering microfluidic device of one of claims 6 and 7, wherein an outlet of a primary connection (41 , 42, 43) into to the meeting and merging cavity (32) is configured to hold a flow of liquid to be mixed within the control volume connected to this primary connection.

9. The metering microfluidic device of any preceding claim, wherein the meeting and merging cavity (32) is defined between at least two opposite walls (322, 324); an outlet of any primary connection (41 , 42) but the last one (43) is located on a first wall (322) among the two opposite walls and configured so that the liquid to be mixed coming out of the primary connection adheres on the first wall (322) without contacting the second wall (324); and the outlet of the last primary connection (43) is configured so that the last liquid to be mixed contacts the two opposite walls (322, 324) when coming out of the last primary connection.

10. The metering microfluidic device of claim 9, wherein a downstream outlet of any primary connection (41 , 42) but the last one (43) is of the curve-curve type (b), of the right-curve type (c) or of overhanging curve-curve type (d), whereas a downstream outlet of the last primary connection (43) is defined by a sharp edge and centered between the two opposite walls (322, 324) of the meeting and merging cavity (32).

11. The microfluidic device of any preceding claim, wherein it has no valve for controlling the flow of liquids to be mixed, into and out of the control volumes (Vc1 , Vc2, Vc3).

12. The metering microfluidic device of any preceding claim, wherein it is formed by a stack of plates (22, 24, 26) which define parts (R12, R22, R32, R14, R24, R34) of the overflow reservoirs (R1 , R2, R3), parts of the control volumes (Vc1 , Vc2, Vc3) and / or parts (i12, i22, i32, i14, i24, i34, i16, i26, i36, e12, e22, e32, e14, e24, e34, e16, e26, e36) of the injection or entry ports (i1 , i2, i3, e1 , e2, e3).

13. The metering microfluidic device of any one of claims 1 to 11 , wherein it is monobloc, preferably made by 3D printing, stereo photolithography, injection molding or sintering.

14. A process for mixing at least two liquids in a metering microfluidic device (2) according to any preceding claim, comprising at least the following steps:a) injecting the first liquid (L1) to be mixed into the first control volume (Vc1), via the first injection port (i 1 ) , until a portion of the first liquid enters the first overflow reservoir (R1) via the first secondary connection (51); b) injecting the last liquid (L3) to be merged into the last control volume (Vc3), via the last injection port (i3), until a portion of the last liquid enters the last overflow reservoir (R3) via the last secondary connection (53); c) injecting the first pushing gas (A1) into the first control volume (Vc1), via the first entry port (e1), until all the first liquid to be mixed contained in the first control volume is pushed by the first pushing gas into the meeting and merging cavity (32), via the first primary connection (41); and d) injecting the last pushing gas into the last control volume (Vc3), via the last entry port (e3), until all the last liquid to be mixed contained in the last control volume is pushed by the last pushing gas into the meeting and merging cavity (32), via the last primary connection (43).

15. The process of claim 14, wherein the pushing gas is the same for all control volumes.

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