Method and device for microfluidic manipulation based on thermo-viscous liquid

Abstract

The present invention relates to a microfluidic manipulation device and method comprising the steps: (a) injecting a sample liquid (20) into a common channel (4) of a microfluidic device; (b) injecting at least one sheath liquid (21, 22) comprising a thermo-viscous liquid having a dynamic viscosity of less than 0.1 Pa.s at a determined injection temperature into the common channel (4). According to the invention, the method comprises step (c) applying an energy source for a limited duration, less than or equal to 10 ms, in at least one heating zone (31) located in the common channel (4) upstream of the branch between the first outlet channel (11) and the second outlet channel (12) to locally heat said sheath liquid (21, 22) which has a dynamic viscosity greater than or equal to 1.0 Pa.s at the high temperature Th and to extract a portion (120, 130, 220) of the sample liquid. Figure for abstract: Figure 1

Description

Title of the invention: Method and device for microfluidic manipulation based on thermo-viscous liquid Technical field of the invention

[0001] The present invention relates to the technical field of microfluidic manipulation devices and methods for rapidly controlling and diverting the flow direction of a liquid. The present invention finds applications in the precise and rapid manipulation of liquids or the extraction of very small volumes within a liquid. The invention particularly relates to the manipulation and sorting of individual nano-sized particles in a liquid. State of the art

[0002] In the above field, it is known to use flow cytometers to detect, count, identify cells or particles of micrometric dimensions suspended in a flowing liquid by passing them, one by one and at high speed, in the beam of one or more lasers and then to sort them by implementing sorting methods based on various techniques.

[0003] Advances in microscopy and biology have made it possible to focus on objects of ever smaller size, in particular nano-objects of size less than 100 nm.

[0004] Traditional flow cytometers only work properly for objects larger than a micrometer. For nanometric objects or nanoobjects, i.e. smaller than 100 nm, cytometers are currently limited by their detection capacity. In addition, regarding the operation of sorting nano-objects, the volumes of liquid generally used by known cell sorters and the size of the drops used to encapsulate the objects to be sorted are not compatible with the sorting of nanometric objects.

[0005] There are different methods for sorting objects in a microfluidic chip. Some sorting methods rely on passive techniques. They can be based on inertial focusing methods or on the structure of the chip itself: this allows objects to be separated into subpopulations based on their physical characteristics (volume, mass, etc.). Other sorting methods, also passive, rely on the microfabrication of micrometric pillars and allow on-chip chromatography to be carried out, but are only applicable for micrometric objects or polymer chains of micrometric length and polydisperse. These passive sorting methods do not allow objects to be sorted one by one based on a specific signal, for example fluorescence.

[0006] Micromechanical techniques allow sorting of micro-sized objects metric at a relatively slow sorting rate. Thus, the use of valves to separate cells, for example eukaryotic or prokaryotic cells, does not allow a sorting rate of 1 Hz to be exceeded.

[0007] Other active sorting techniques rely on the application of an electric or magnetic field. For example, dielectrophoresis or the use of surface acoustic waves make it possible to achieve a sorting rate of the order of kHz on preformed micrometric drops encapsulating individual objects.

[0008] The microbiological analysis of nanometric objects of interest is all the more difficult since these nanometric objects of interest are present in a sample liquid generally containing a large number of other objects of all kinds from which the nanometric objects of interest are to be isolated. To take only the nano-objects of interest, it is therefore necessary to extract very small volumes of sample liquid each time one wishes to extract an object of interest isolated from the other objects. In addition, the same sample liquid may contain different types of nanometric objects that one wishes to separate from each other. Furthermore, it is desirable to increase the concentration of nanometric objects of a given type in a liquid in order to be able to analyze these nanometric objects. Given the very small size of the nanometric objects, it is necessary to extract a sufficient quantity to carry out a subsequent analysis, for example proteomics.It is currently estimated that approximately 108 nanometric objects of interest are required to perform a proteomic analysis. To sort this quantity of nano-objects in a reasonable time, for example of the order of 3 hours, it is necessary to be able to sort them at a rate of approximately 104 nano-objects per second, or 10 kHz. Thus, the sorting frequency is an essential criterion for the sorting of nano-objects to be useful.

[0009] An aim of the present disclosure is to propose a microfluidic manipulation device and method capable of sorting nanometric objects in a liquid at a high sorting frequency, for example greater than 1kHz and preferably at least 10 kHz, without risk of deterioration of the nanometric objects thus manipulated.

[0010] One of the aims of the present disclosure is to provide a microfluidic manipulation device and method having a faster time response, to enable the extraction of a very small volume of a sample liquid, between a few femtoliters and a few hundred femtoliters or to enable the sorting of individual objects at a higher sorting frequency than what is available in the prior art.

[0011] Another aim of the present disclosure is to propose a microfluidic manipulation device and method for selectively extracting nanometric objects from an initial sample liquid to obtain a new liquid sample in which the concentration of these nanometric objects is higher than that in the initial sample liquid and / or in which the number of these nanometric objects is high enough, for example above 108.

[0012] Yet another object of the present disclosure is to provide a microfluidic manipulation device and method for rapidly obtaining a sample liquid concentrated in selected nanometric objects. Presentation of the invention

[0013] In order to overcome the aforementioned drawbacks of the state of the art, the present invention proposes a microfluidic manipulation method comprising the following steps: (a) injecting via an inlet channel a sample liquid into a common channel of a microfluidic device; (b) injecting via at least one other inlet channel at least one sheath liquid at a determined injection temperature into the common channel to allow hydrodynamic focusing of the sample liquid in the common channel and flow of the sample liquid towards a first outlet channel which is located downstream of a branch between the first outlet channel and a second outlet channel, the at least one sheath liquid comprising a thermoviscous liquid, said at least one sheath liquid comprising the thermoviscous liquid having a dynamic viscosity of less than 0.1 Pa.s at the injection temperature.

[0014] According to the invention, the method comprises the following step: (c) applying an energy source for a limited duration, less than or equal to 10 ms, preferably less than 5 ms, 1 ms, 500 ps or even 1 ps, in a heating zone located in the common channel upstream of the branch between the first outlet channel and the second outlet channel, the heating zone being located in or in contact with the at least one sheath liquid comprising the thermo-viscous liquid so as to locally heat said at least one sheath liquid to a high temperature Th in the heating zone of the common channel, the injection temperature and the high temperature being separated by a temperature difference less than or equal to 40°C, said at least one sheath liquid comprising the thermo-viscous liquid having a dynamic viscosity greater than or equal to 1.0 Pa.s at the high temperature Th, so as to deflect or extract a portion of the sample liquid selectively towards the second outlet channel.

[0015] Advantageously, the at least one sheath liquid comprising the thermoviscous liquid has, following localized heating in the heating zone, a thermal variation in viscosity between the injection temperature and the high temperature, adapted to deflect or extract a portion of the sample liquid selectively towards a determined outlet channel among the at least two outlet channels.

[0016] The method is configured to avoid generating bubble(s) and heating the sample liquid and to avoid heating the sheath liquid over its entire cross-section. Remarkably, the brief and local heating of the sheath liquid does not cause any blockage of the flow either in the common channel or on any of the outlet branches but simply a limited disturbance in time and space of the flow and its distribution in the common channel. This disturbance causes a dynamic modification of the flow downstream of the heating zone, without interrupting the flow, which makes it possible to obtain a rapid and localized effect of deflection of the sample liquid, unlike the prior art.

[0017] Due to the pointed shape of the branch between the two outlet channels, a disturbance in the viscosity of the sheath liquid causes sufficient deflection of the sample liquid to deflect it towards the first outlet channel. As soon as the heating is interrupted, the heated and viscous zone is carried away in the flow and quickly returns to the injection temperature.

[0018] Advantageously, the injection temperature and the high temperature are separated by a temperature difference less than or equal to 40°C, 30°C, 20°C or even 10°C.

[0019] Such a method makes it possible to divert or extract a portion of the sample liquid having a very small volume, between one femtoliter and 900 femtoliters, for example of the order of 10 to 100 femtoliters. This method allows the manipulation and sorting of individual cells or particles of nanometric size suspended in the sample liquid. In addition, the method allows sorting at a higher rate than in the methods of the prior art.

[0020] According to a particular and advantageous aspect, the thermoviscous sheath liquid comprises a thermosensitive polymer.

[0021] Advantageously, the sheath liquid comprises from 1% to 30% by weight of said heat-sensitive polymer.

[0022] In one example, the heat-sensitive polymer comprises at least one linear chain of heat-sensitive polyoxyalkylene type extended at at least one of its ends by an organic group via a carbamate or ester bond.

[0023] In another example, the heat-sensitive polymer comprises, on the one hand, water-soluble units and, on the other hand, units chosen from polyurethanes comprising poly(ethylene oxide-b-propylene oxide-b-ethylene oxide) (POE-b-POP-b-POE) groups and being present at a mass concentration of less than or equal to 10% in an aqueous solution.

[0024] According to a first embodiment, the method comprises a following step after step (c):

[0025] (d) applying another energy source for a limited duration, less than or equal to 10 ms, in a second heating zone located in the common channel upstream of the branch between the first outlet channel and the second outlet channel, the second heating zone being located in or in contact with the at least one sheath liquid comprising the thermo-viscous liquid to locally heat said at least one sheath liquid to the high temperature Th in the second heating zone of the common channel so as to restore the flow of the sample liquid towards the first outlet channel.

[0026] Advantageously, step d) is separated from step c) by a duration less than or equal to 500 ps.

[0027] According to a particular and advantageous aspect, the energy source of step c) comprises a laser emitting a first laser pulse at a wavelength between 100 nm and 10 pm.

[0028] According to another particular aspect, the other energy source of step d) comprises a laser emitting a second laser pulse at a wavelength between 100 nm and 10 pm.

[0029] Advantageously, the sheath liquid comprising the thermo-viscous liquid is capable of absorbing the first laser pulse and / or the second laser pulse.

[0030] For example, the sheath liquid comprises a component in suspension or in solution capable of absorbing the first and / or the second laser pulse.

[0031] According to another particular and advantageous aspect, the heating zone or, respectively, the second heating zone comprises a photo-thermal transducer.

[0032] According to yet another particular and advantageous aspect, the first heating zone or, respectively, the second heating zone, comprises an electrothermal transducer and in which the energy source of step c), respectively of step d), comprises an electrical energy source.

[0033] Preferably, the high temperature Th is less than or equal to 60°C, 50°C or 40°C or 35°C or 30°C.

[0034] The at least one sheath liquid has a dynamic viscosity that increases as a function of temperature in a determined temperature range greater than or equal to the injection temperature.

[0035] The determined temperature range extends, above the injection temperature, over approximately 10 degrees, or 15 degrees, 20 degrees, 25 degrees or even 30 degrees, 35 degrees or 40 degrees.

[0036] According to a particular aspect, the thermal variation of dynamic viscosity has a maximum greater than or equal to 3 Pa.sK A or 5 Pa.sK *, or even 7 Pa.sK 1 in the determined temperature range.

[0037] Advantageously, the sample liquid is free of thermoviscous liquid.

[0038] The invention also relates to a microfluidic device comprising a sample liquid inlet channel, at least one other sheath liquid inlet channel, a first outlet channel, a second outlet channel, a common channel arranged between said inlet channels and said outlet channels, the common channel being fluidically connected to said inlet and outlet channels, a branch separating downstream the first outlet channel from the second outlet channel, the sample liquid inlet channel containing a sample liquid injected into the common channel, said at least one other sheath liquid inlet channel containing at least one sheath liquid injected into the common channel at a determined injection temperature, the at least one sheath liquid comprising a thermo-viscous liquid, said at least one sheath liquid comprising the thermo-viscous liquid having a dynamic viscosity of less than 0.1 Pa.s at the injection temperature, so as to allow hydrodynamic focusing of the sample liquid in the common channel, the microfluidic device being configured to conduct the sample liquid hydrodynamically focused in the common channel by the at least one sheath liquid at the injection temperature towards the first outlet channel.

[0039] According to the invention, the microfluidic device comprises heating means comprising an energy source and at least one heating zone located in the common channel upstream of the branch between the first outlet channel and the second outlet channel, said at least one heating zone being located in or in contact with the at least one sheath liquid comprising the thermoviscous liquid, the heating means being configured to apply the energy source for a limited duration, less than or equal to 10 ms, in said at least one heating zone, so as to locally heat said at least one sheath liquid to a high temperature Th in the common channel in said at least one heating zone of the common channel, the injection temperature and the high temperature being separated by a temperature difference less than or equal to 40°C, and preferably less than or equal to 30°C, 20°C or even 10°C,and said at least one sheath liquid comprising the thermo-viscous liquid having a dynamic viscosity greater than or equal to 1.0 Pa.s at the high temperature Th, so as to deflect or extract a portion of the sample liquid selectively towards the second outlet channel.

[0040] According to a particular and advantageous aspect, the energy source comprises a pulsed laser capable of generating pulses having an energy per pulse of between 100 nJ and 1000 nJ, for example of the order of 100 nJ.

[0041] Of course, the various features, variants and embodiments of the invention may be combined with each other in various combinations to the extent that they are not incompatible or mutually exclusive. Detailed description of the invention

[0042] Furthermore, various other characteristics of the invention emerge from the appended description given with reference to the drawings which illustrate a non-limiting form of carrying out the invention and where:

[0043] [Fig.l] is a schematic view of a microfluidic device and method according to the present disclosure;

[0044] [Fig.2] represents a solid line curve of viscosity as a function of temperature and respectively a dashed curve of viscosity variation as a function of temperature for an example of sheath liquid based on thermoviscous liquid;

[0045] [Fig.3] illustrates a first embodiment of the method based on the application of two successive energy pulses causing a rise in temperature of the sheath liquid, with here the application of the first pulse;

[0046] [Fig.4] illustrates the first embodiment, just after the application of the second pulse;

[0047] [Fig.5] illustrates the first embodiment and the extraction of a portion of sample liquid to an outlet channel;

[0048] [Fig.6] illustrates the first embodiment and the extraction of a portion of sample liquid to an outlet channel;

[0049] [Fig.7] illustrates a second embodiment of the method based on the application of a single pulse, during this pulse;

[0050] [Fig.8] illustrates the second embodiment after the end of this pulse and the extraction of a portion of sample liquid towards an outlet channel.

[0051] In this document, the term thermo-viscous liquid means a liquid whose viscosity increases strongly and reversibly with temperature.

[0052] [Fig. 1] represents a microfluidic device 100 according to an exemplary embodiment of the present disclosure. The microfluidic device 100 is generally a planar device commonly called a microfluidic chip. An orthonormal XY reference frame is represented in the plane of [Fig. 1].

[0053] The microfluidic device 100 comprises inlet channels 1, 2 and 3, a common channel 4 and at least two outlet channels 11, 12. The inlet channels 1, 2 are fluidically connected respectively to a source of sheath liquid, which is for example stored in a common reservoir or in two separate reservoirs. The inlet channel 3 is fluidically connected to a source of sample liquid 20 to be analyzed, which is for example stored in another reservoir. At the other end of the common channel 4 is a junction, for example of the Y junction type comprising at least a first outlet channel 11 and a second outlet channel 12 generally arranged symmetrically with respect to the longitudinal axis 14 of the common channel 4. The longitudinal axis 14 is here parallel to the X axis of the XY reference frame. A branch, here in the form of a point, separates the first outlet channel 11 from the second outlet channel 12. The point of the branch forms an acute angle generally between 5 deg.and 40 deg. In another variant, the microfluidic device has three output channels: one . first outlet channel in the longitudinal axis 14 of the common channel, a second outlet channel disposed on one side of the common channel and a third outlet channel disposed on an opposite side of the common channel. In another variant, the microfluidic device comprises more than three outlet channels. In these variants, a branch separates each pair of adjacent outlet channels.

[0054] In a known manner, the microfluidic device 100 is for example manufactured from glass, ceramic or silicon or a polymer such as polydimethylsiloxane (PDMS). The microfluidic device can be manufactured monolithically or by assembling a support block and a cover-forming slide. The support is for example manufactured from PDMS. The channels 1, 2, 3, 4, 11, 12 are formed for example by molding during the manufacture of the support. The support has for example the following dimensions: 5 mm thick and 20 to 40 mm on each side. The cover-forming slide is preferably transparent to allow observation and detection of particles in the sample liquid. For example, the slide is a microscope slide to allow observation of the microfluidic device 100 under an optical microscope objective. For example, a very thin glass slide is used, having a thickness of 0.15 mm.

[0055] We denote W the width, H the depth and L the length of the common channel 4. For example, in a microfluidic device as illustrated in [Fig.l], the width W of the common channel is 25 pm, the depth H of the common channel is 7 pm and the length L is approximately 100 pm. The distance G between the start of the outlet channel 11, 12 and the tip of the branch of the junction Y is between 10 and 30 pm.

[0056] According to one variant, the microfluidic device has a symmetry of revolution around a longitudinal axis. In this case, a single sheath liquid inlet channel can be used.

[0057] The sample liquid 20 is composed, for example, of water or a phosphate buffer saline (PBS) solution in which micro-objects or nano-objects to be categorized and manipulated are suspended. The sample liquid 20 comprises, for example, particles to be sorted, extracted and / or concentrated.

[0058] The sample liquid 20 and the sheath liquid(s) 21, 22 may be miscible or immiscible with each other. In general, the first sheath liquid 21 and the second sheath liquid 22 are identical. Advantageously in this case, the first sheath liquid 21 and the second sheath liquid 22 are stored in a single source reservoir. Either motorized syringes (commonly called "syringe pumps") are used in which the flow rate of the syringes is controlled, or systems based on pressure generators in which the injection pressure is controlled to inject the sample liquid 20 and respectively the sheath liquid(s) 21, 22 into the microfluidic device 100.

[0059] A first sheath liquid 21 is injected simultaneously via the inlet channel 1, a second sheath liquid 22 via the inlet channel 2 and the sample liquid 20 via the inlet channel 3. The first sheath liquid 21 and / or the second sheath liquid 22 are injected at a determined injection temperature into the common channel 4. For example, the injection temperature is equal to the ambient temperature of 18°C, 19°C, 20°C or 22°C or 25°C.

[0060] The inlet channels 3, 1, 2 and fluidly conduct the sample liquid 20 to be analyzed and the sheath liquid(s) 21, 22 to the common channel 4 where the sample liquid 20 is hydrodynamically focused. Depending on the ratio between the flow rate of the sample liquid and the total flow rate of the sheath liquids, the sample liquid 20 has, in the common channel 4, a width D reduced compared to the width W by hydrodynamic focusing. For example, a sample liquid stream 20 of reduced width D of between 2 and 7 micrometers is thus obtained. At the outlet of the common channel, and when no sorting action is in progress, the sample liquid stream 20 is continuously directed towards one or other of the outlet channels 11, 12.

[0061] The microfluidic device 100 comprises a detection unit 50 arranged in the common channel 4. The detection unit 50 is for example based on a system for detecting one or more fluorescence signals emitted by particles passing in the common channel 4, these particles to be detected being marked by one or more fluorescent markers. The particles with fluorescent marker are for example particles included in the sample liquid, which it is desired to sort or extract. For example, the fluorescence is measured at one or more wavelengths and the sorting is activated when a fluorescence signal comprising a predetermined combination of these wavelengths is detected. The detection unit 50 can also be based on the measurement of particle size by scattering one or more laser beams incident on the particle, or by combining several optical or electrical methods.

[0062] The microfluidic device 100 comprises at least a first heating zone 31 located in the common channel 4, downstream of the detection unit 50. By way of example, in [Fig. 1], two heating zones 31, 32 are shown located in the common channel 4, upstream of the outlet channels 11, 12. The first heating zone 31 is located on the same side of the microfluidic device 100 as the inlet channel 1 and the first outlet channel 11. The second heating zone 32 is located on the same side of the microfluidic device 100 as the inlet channel 2 and the second outlet channel 12. A laser beam 41, respectively 42, focused in the first heating zone 31, respectively the second heating zone 32, makes it possible to locally heat the sheath liquid 21, respectively 22. In the plane of the [Fig.l], each heating zone 31, 32 extends over a limited surface, by example in the shape of a disc, square or rectangle. In particular, each heating zone 31, 32 is limited transversely, along the Y axis of the XY reference frame, to a width less than the width of the first sheath liquid 21, respectively of the second sheath liquid 22. Advantageously, each heating zone 31, 32 has a width less than or equal to 35%, 25%, 20% or 10% of the width W. In the example above, the width W is 25 μm and the width of each heating zone 31, 32 is approximately 6 μm.

[0063] The first heating zone 31, respectively the second heating zone 32, is located at a distance D1, respectively D2, from the detection unit 50. The first heating zone 31, respectively the second heating zone 32, is located at a distance PI, respectively P2, from the tip of the branch of the Y junction which separates the first output channel 11 from the second output channel 12. By way of example, the distance D1, respectively D2, is between 1 and 10 pm and the distance PI, respectively P2 is between 0 and 20 pm.

[0064] According to a variant, the two heating zones 31, 32 are located in the common channel 4 on the same side of the microfluidic device 100 as the inlet channel 1 and the first outlet channel 11. In this case, the first heating zone 31 is located upstream of the second heating zone 32.

[0065] The microfluidic device 100 is generally of planar shape. However, in another embodiment, the inlet channel 3 of the sample liquid 20, the common channel 4, the first outlet channel 11 and the parts connected to the sheath liquid inlet channel 1 and to the second outlet channel 12 are of revolution around the longitudinal axis 14. Alternatively, it is possible to combine rotationally symmetrical inlet channels with a common channel and outlet channels of planar geometry. In this variant, the microfluidic device 100 comprises a single sheath fluid inlet channel or two or more sheath fluid inlet channels. According to another variant, it is possible to combine inlet channels and a common channel of planar geometry with rotationally symmetrical outlet channels.

[0066] According to the present disclosure, a sheath liquid having particular properties of viscosity variation as a function of temperature is used. More precisely, at least one sheath liquid 21 and / or 22 comprising a thermoviscous liquid is used.

[0067] The viscosity of the thermoviscous liquid increases sharply and reversibly with temperature. The thermoviscous liquid preferably has an initial viscosity close to that of water at the injection temperature, for example at room temperature, and its viscosity increases as the temperature increases. Conversely, when the temperature of the thermoviscous liquid decreases and returns to room temperature, the thermoviscous liquid sees its viscosity decrease for return to its initial value.

[0068] The sheath liquid 21, 22 comprises, for example, a thermoreversible gel of the hydrogel type based on a mixture of water and a thermosensitive polymer or thermosensitive polymer. A thermosensitive polymer is, for example, made up of thermosensitive hydrophobic segments of the polypropylene oxide (POP) type and hydrophilic parts of the polyethylene oxide (PEO) type. Such thermosensitive polymers are used in cosmetics.

[0069] In one example, the heat-sensitive polymer comprises at least one linear chain of heat-sensitive polyoxyalkylene type extended at at least one of its ends by an organic group via a carbamate or ester bond as described in patent document FR2840907 from the company Polymerexpert for applications in the field of cosmetics.

[0070] In another example, the heat-sensitive polymer comprises, on the one hand, water-soluble units and, on the other hand, units chosen from polyurethanes comprising poly(ethylene oxide-b-propylene oxide-b-ethylene oxide) (POE-b-POP-b-POE) groups and being present at a mass concentration of less than or equal to 10% in an aqueous solution as described in patent document FR2940761 of the company Polymerexpert for applications to a formulation for the treatment and / or prevention of snoring and administrable by spraying via the nasal or oral routes.

[0071] The thermo-viscous liquid is manufactured for example by dissolving a thermo-sensitive polymer in an aqueous solution, for example water, to which one or more additives are optionally added to adjust the viscosity properties of the thermo-viscous liquid.

[0072] For example, a sheath liquid comprising 5% by weight of thermosensitive polymer diluted in pure water is used. Advantageously, the sheath liquid comprises approximately 2% by weight of a substance exhibiting strong absorption at the laser wavelength, for example ink, or dye, or pigment, to enable absorption of a laser beam. Preferably, a biocompatible, food-grade black ink is used. Alternatively, carbon nanoparticles are suspended in the sheath liquid, for example nanoparticles having a size of approximately 90nm.

[0073] These families of thermo-viscous liquids are characterized by a low viscosity at room temperature, the viscosity being less than or equal to 100 mPa.s, or even 40 mPa.s, for example between 10 mPa.s and 30 mPa.s, (or even less than 10 mPa.s in certain physicochemical configurations), and an increasing viscosity in a determined temperature range greater than or equal to room temperature until reaching a maximum viscosity of 1.0 Pa.s or several Pa.s or even several tens of Pa.s, at a temperature of the order of 37°C + / - 10°C. This results in a variation in viscosity as a function of temperature, the maximum of which can reach 1 Pa.s / K or several Pa.s / K, for example 5 Pa.s / K or 10 Pa.s / K, in a reduced temperature range from room temperature.

[0074] The minimum and maximum viscosity values ​​and the thermal viscosity variation values ​​as well as the temperature corresponding to the maximum viscosity value depend on the thermo-viscous liquid chosen, its mass concentration in water and possibly the additive(s) incorporated in the sheath liquid.

[0075] The thermo-viscous liquid has a dynamic viscosity which increases as a function of temperature in a determined temperature range from an injection temperature into the microfluidic chip. The injection temperature is between 5°C and 30°C, preferably between 15°C and 25°C, preferably between 18°C ​​and 22°C. Advantageously, the injection temperature is equal to the ambient temperature of 18°C, 19°C, 20°C, 21°C, 22°C, 23°C, 24°C or 25°C.

[0076] For example, a thermo-viscous sheath liquid 21, 22 is used comprising a thermosensitive polymer from the company PolymerExpert with the trade name EG230 (CAS number 93665-35-1) which is a branched polyurethane containing POE-b-POP-b-POE units, the INCI (International Nomenclature of Cosmetic Ingredients) designation of which is BISMethoxy PEG-13 PEG-502 / PPG-57 SMDI Copolymer. The percentage by weight of thermosensitive polymer in the sheath liquid is between 0.5% and 10%, preferably between 1% and 7.5%, for example 5%, 6% or even 7.5%, the thermosensitive polymer being diluted in pure water.

[0077] [Fig.2] represents a viscosity curve V (solid line) as a function of temperature and respectively a viscosity variation curve AV / AT (dashed line) as a function of temperature, between 12°C and 50°C, for a sheath liquid based on thermosensitive polymer EG230 diluted to a concentration of 7.5% in water. It is observed that the viscosity V of the sheath liquid comprising this thermo-viscous liquid increases continuously with the temperature between 12°C and approximately 45°C unlike usual sheath liquids. In addition, the viscosity value below 22°C is very low: of the order of 0.02 Pa.s. Finally, the maximum viscosity value is very high in this example: this maximum value reaches approximately 35 Pa.s at the temperature of 46°C. Above the temperature of 46°C, the viscosity decreases continuously while remaining at high values, for example about 28 Pa.s at 50°C.

[0078] It is observed that the slope of thermal variation of the viscosity of the sheath liquid comprising the thermo-viscous liquid based on thermosensitive polymer is positive and particularly strong over a small temperature range between approximately 25°C and 40°C. This slope is approximately 0.3 Pa.s / K around 30°C, approximately 1.0 Pa.s / K around of 33°C, of ​​about 2.0 Pa.s / K around 36°C and of about 4.0 Pa.s / K around 40°C. In other words, the maximum slope of the thermal viscosity variation curve in this example is 4.0 Pa.s / K. Above 46°C, the thermal viscosity variation slope of the sheath liquid including the thermoviscous liquid becomes negative.

[0079] In another example, a sheath liquid comprising a heat-sensitive polymer diluted in pure water is used. The sheath liquid has a very low viscosity of less than or equal to 0.04 mPa.s at a temperature of less than or equal to 20°C, and a strong increase in its viscosity between 20°C and 40°C, with a positive slope exceeding 10 Pa.s / K. At a temperature of 35°C, the sheath liquid has a viscosity of approximately 5000 mPa.s. At a temperature of 40°C, the sheath liquid has a viscosity of approximately 18000 mPa.s. Above 40°C, the sheath liquid has a viscosity which decreases with temperature.

[0080] It is observed that a small increase in temperature of the order of 10°C, 15°C, 20°C or 30°C, relative to the ambient temperature of approximately 20°C, is sufficient to significantly increase the viscosity of the sheath liquid based on thermoviscous liquid.

[0081] In addition, this sheath liquid is thermoreversible. Consequently, this viscosity variation is reversible. A decrease in temperature of the thermoviscous sheath liquid based on the thermosensitive polymer EG230 7.5% from 40°C to 20°C makes it possible to reduce the viscosity from a value equal to approximately 20 Pa.s (at 40°C) to a value less than or equal to 0.04 mPa.s (at 20°C).

[0082] According to the present disclosure, the sample liquid generally consists of cells or particles of nanometric size suspended in a generally water-based liquid. The sample liquid consists of a medium suitable for the transported particles, which may be pure water, or a phosphate buffered saline solution (or PBS, from the English phosphate-buffered saline) or even a nutrient medium for biological particles. The sample liquid has a dynamic viscosity which slightly decreases as a function of the temperature between 0°C and 100°C and in particular in the determined temperature range from the injection temperature, which is preferably room temperature. The sample liquid 20 is free of thermo-viscous liquid. Indeed, the addition of thermo-viscous liquid to this sample liquid is likely to harm the integrity of the particles which it is sought to isolate.

[0083] The microfluidic device takes advantage of the properties of at least one thermo-viscous sheath liquid 21 and / or 22 to modify the flow of the sample liquid 20 in order to carry out sorting or extraction of a portion of sample liquid towards a dedicated outlet channel.

[0084] We will now explain the operation of the microfluidic device in link with figures 1 and 3-8.

[0085] The sample liquid 20 is injected into the microfluidic chip 100 via the inlet channel 3. The sheath liquids 21 and 22 respectively are injected into the microfluidic chip via the inlet channels 1 and 2 respectively. In the example illustrated, the microfluidic chip 100 has, at the level of the common channel 4, a depth of 7 μm and a width of 25 μm, taken perpendicular to the longitudinal direction of propagation of the liquids. The sheath liquids 21, 22 are identical here. According to the present disclosure, the injection temperature of the sheath liquid is approximately equal to ambient temperature, for example 20°C. The flow rate of the sample liquid 20 is for example 5 μl / h.In the illustrated examples, the respective flow rates of the two sheath liquids 21, 22 are adjusted to slightly different values ​​(for example 80 pl / h and 81 pl / h respectively) to direct the sample liquid 20 towards the first outlet channel 11 in the absence of heating in the heating zones. In this configuration, the first outlet channel 11 is a so-called trash outlet and the second outlet arm 12 is used to collect the sorted particles.

[0086] At room temperature, the low viscosity of the thermo-viscous liquid-based sheath liquid makes it possible to produce flows having a high linear speed (several tens of cm / s) despite the small dimensions of the channels of the microfluidic chip. In addition, the very strong reversible variation in viscosity as a function of temperature allows rapid switching by means of a small rise and then fall in temperature in the sheath liquid.

[0087] In an exemplary embodiment, the energy source used is a laser emitting a laser beam at a determined wavelength and the sheath liquid 21, 22 is capable of absorbing the laser beam to transform it into heat. More precisely, a laser beam 41 is directed and focused in the first heating zone 31 with a beam size of between approximately 1 μm and 10 μm in the focusing zone. In a complementary and optional manner, another laser beam 42 is directed and focused in the second heating zone 32, as illustrated in [Fig.4].

[0088] For example, the sheath liquid 21, 22 is composed of a thermo-viscous liquid based on a thermosensitive polymer dissolved in water and a material absorbing at the wavelength of the laser. The absorbing material is in suspension or solution in the sheath liquid. The absorbing material comprises for example an ink or carbon nanoparticles, capable of absorbing the laser beam focused inside the sheath liquid in the heating zone 31, 32 to transform it into heat. Alternatively, the wavelength of the laser beam corresponds to an absorption line of the liquid carrying the sheath liquid, for example an absorption line of water. The heating zone 31, 32 has a diameter of approximately 5 μm in [Fig.l].

[0089] In another exemplary embodiment, each heating zone 31, 32 comprises a photothermal transducer made in such a way as to be in contact with the sheath liquid, each photothermal transducer being capable of absorbing the laser beam focused in said heating zone 31, 32. Each photothermal transducer comprises a metallic surface, formed by deposition of a thin metallic layer, for example gold or indium, with a thickness of between 10 nm and 500 nm and having an area of ​​between 1 pm2 and 100 pm2. Each photothermal transducer is capable of absorbing the laser beam focused in the heating zone 31, 32 to transform it into heat locally.

[0090] According to yet another exemplary embodiment, each heating zone 31, 32 comprises an electro-thermal transducer, for example a heating resistor or an inductive dissipative element. Each electro-thermal transducer is connected to a source of electrical energy. The electro-thermal transducer is in contact with the first, respectively second, sheath liquid 21, 22 in the common channel 4 to allow a localized heat exchange.

[0091] [Fig.l] illustrates an example in which a laser beam 41, respectively 42, directly heats the sheath liquid 21, respectively 22, in the first heating zone 31, respectively the second heating zone 32. The sheath liquid 1, 2 is composed of thermoviscous liquid and a material absorbing at the wavelength of the laser, in suspension or solution in the sheath liquid. The thermoviscous liquid is composed of water and a thermosensitive polymer, here EG230 with a mass concentration of 7.5% in water. The sample liquid 20 consists of water containing the particles of interest.

[0092] The heating zone is located in or in contact with the sheath liquid and preferably at a non-zero distance from the sample liquid. For example, the distance between the first heating zone 31, 32 and the sample liquid 20 is of the order of 4 μm. In this way, the temperature increase is precisely located inside the sheath liquid comprising the thermoviscous liquid. This arrangement makes it possible to spatially limit the temperature rise in the sheath liquid and outside the sample liquid, so as to avoid a temperature rise in the sample liquid likely to damage or modify the particles located in the sample liquid. In addition, the temperature rise is spatially limited to a portion of the cross-section of the sheath liquid so as to avoid blockage of the common channel 4 or one of the outlet channels 11, 12 by the sheath liquid when its viscosity increases.

[0093] In a first exemplary embodiment illustrated at different times in FIGS. 3 to 6, following the detection via the detection unit 50 of a particle of interest in the sample liquid, the emission of a sequence of two successive laser pulses 41, 42 is triggered, each having a limited duration or short duration, between 1 ps and 500 ps, ​​for example here of the order of 50ps. The laser power is of the order of 5mW. The energy per pulse is between 10000 nJ and 10000 nJ, for example here 250 nJ. The first laser pulse 41 is focused in the first heating zone 31 for a duration of 50ps ([Fig.3]) then, after an interval of duration between 0 and 50ps, the second laser pulse 42 is focused in the second heating zone 32 for a duration of 50ps ([Fig.4]). It is assumed that the sheath liquid absorbs all of the power of the laser pulses.

[0094] Figures 3 to 6 illustrate the two-laser pulse sequence well. In [Fig. 3], the first laser pulse 41 in the first heating zone 31, on a first side of the microfluidic device 100, initiates the deflection of the sample liquid toward the second outlet channel 12. In [Fig. 4], as soon as the deflection is initiated, the second laser pulse 42 in the second heating zone 32, on a second side of the microfluidic device 100, reverses the movement. However, since the sample liquid has propagated between the two laser pulses, a small portion of the sample liquid continues toward the second outlet channel 12. In [Fig.5], the two laser pulses 41, 42 are extinguished, but the effect of local increase in the viscosity of the sheath liquid continues to propagate: an extracted portion 130 is observed forming in the second outlet channel 12 while the remainder of the sample liquid flow 20 returns to the first outlet channel 11 or waste arm. In [Fig.6], the extracted portion 130 has detached from the sample liquid flow 20 which is restored in the first outlet channel 11 (waste arm).

[0095] Figures 3-8 also show the local temperature change of the sheath liquid 21, 22 during and immediately after each laser pulse. The color of the sheath liquid is shown schematically as a function of temperature. The color of the sheath liquid is lighter where the sheath liquid is at room temperature, i.e. where the sheath liquid has a low viscosity, less than 0.1 Pa.s. The color of the sheath liquid is darker locally, the higher the temperature. In other words, the sheath liquid has a viscosity locally that is higher the darker its color in Figures 3-8.

[0096] In Figures 3-8, upstream of the heating zones 31, 32, the sample liquid 20 and the sheath liquid 21, 22 are at the injection temperature, for example at room temperature of approximately 20°C. The injection temperature here is a temperature at which the sheath liquid 21, 22 has a very low viscosity. At 20°C, the sheath liquid 21, 22 comprising the thermo-viscous liquid has a low viscosity of approximately 20 mPa.s. The absorption of each laser pulse 41, respectively 42 locally increases the temperature of the sheath liquid 21, respectively 22 in the first heating zone 31, respectively the second heating zone 32 up to a high temperature Th of approximately 40°C. In the heating zone of the sheath liquid, the temperature increases during the duration of the pulse, it is maximum at the end of the pulse and then decreases at practically the same speed. The limitation in duration and power of the energy input makes it possible to limit the maximum heating temperature of the sheath liquid, here called high temperature Th. This high temperature Th remains well below the boiling temperature of the sheath liquid, so as to avoid the generation of bubbles. The high temperature Th can be lower, equal to or even higher than the temperature at which the sheath liquid has a maximum viscosity value. Advantageously, the high temperature Th is lower than the temperature at which the sheath liquid has a maximum viscosity value, so as to limit heating of the sheath liquid.In addition, the high temperature Th of the sheath liquid is adjusted so that the dynamic viscosity of the sheath liquid at this high temperature Th is significantly higher than the viscosity at the injection temperature. For example, the dynamic viscosity of the sheath liquid is greater than or equal to 1.0 Pa.s at the high temperature Th. This localized and instantaneous heating of the sheath liquid has the effect of making the thermo-viscous liquid extremely viscous in the heating zone during the laser pulse, which causes a deflection of all the streamlines by a modification of the flow conditions.

[0097] Indeed, at 40°C, the thermoviscous liquid has a viscosity greater than 10 Pa.s. This causes a deviation of the flow of the sample liquid stream 20 around the zone of increased viscosity, and in particular a deviation of the sample liquid flow. Stopping the heating causes an almost instantaneous cooling of the sheath liquid, and a reduction in the viscosity of the sheath liquid.

[0098] The spatial limitation of the viscosity increase to a section smaller than the section of the sheath fluid considered makes it possible to avoid the formation of a plug which would block the flow of sheath liquid. This spatial limitation of the viscosity increase inside the sheath liquid makes it possible, on the contrary, to maintain the flow of the sheath liquid towards the outlet channels, which leads to rapid cooling of the sheath liquid after stopping the heating laser pulse. In this way, since the overall flow is not interrupted, the zone of the sheath liquid which has been heated is carried downstream even before it has completely cooled. This further accelerates the restoration of the initial conditions and promotes high-frequency operation.In contrast, in the prior art, the heated zones are located in one of the outlet arms and extend across the entire width of the outlet arm, to produce a plug in the outlet arm, the flow of liquid in this outlet arm then being interrupted as long as the temperature remains high. This operation implies that the restoration of the initial conditions is slow and can only take place after complete cooling of the heated zone.

[0099] According to the present disclosure, the continuous flow of the sheath liquid allows the initial flow direction to be quickly restored after the heating sequence. However, following localized heating of the sheath liquid, the modification of the circulation of the sample liquid 20 makes it possible to direct a very small portion of the sample liquid towards the second outlet channel 12 which serves as a collection path (see Figures 3-6). Nevertheless, the zone of increased viscosity does not block the liquid flow, because it is spatially limited due to the focusing of the laser and it does not extend over the entire width of the sheath liquid. In particular, the liquid flow in the two outlet channels 11, 12 is not interrupted. In addition, the heating time is very short, so that the temperature and viscosity of the zone of increased viscosity decrease very quickly after the heating is stopped.

[0100] In [Fig.6], it is observed that the portion 111 of the sheath liquid having undergone an increase temperature has shifted relative to the position of the first heating zone 31 where the laser pulse 41 was applied. In addition, the maximum temperature of this portion 111 of sheath liquid 21 is approximately 35°C, i.e. lower than the high temperature of 40°C. Indeed, [Fig.4] is carried out 50 ps after the laser pulse 41 has stopped. This shift and rapid reduction in the temperature of the portion 111 of the sheath liquid are explained by thermal diffusion and the continuous flow of the sheath liquid towards the outlet channel 11.

[0101] Similarly, Figures 4 to 6 show the decrease in temperature of the portion 112 of the sheath liquid that has been heated by the second laser pulse 42. In [Fig. 4], the portion 112 of the sheath liquid that has undergone heating is located in the second heating zone 32 and has a temperature of approximately 40°C. In [Fig. 5], the portion 112 of the heated sheath liquid has shifted toward the second outlet channel 12 and has a temperature of approximately 35°C. In [Fig. 6], the portion 112 of the heated sheath liquid has left the second heating zone 32 and is now located in the second outlet channel 12 and has a temperature of approximately 30°C, which corresponds, according to [Fig. 2], to the near restoration of the viscosity at injection temperature.

[0102] Furthermore, it is clearly observed in Figures 3 to 6 that the sample liquid undergoes practically no rise in temperature, in particular in the vicinity of the heating zones 31, 32. Maintaining the sample liquid at the injection temperature makes it possible to preserve the integrity of the particles or cells that it is desired to analyze.

[0103] The application of two successive laser pulses, i.e. offset in time, makes it possible to induce a shearing effect in the sample liquid stream 20, which makes it possible to extract a small portion 130 of the sample liquid in the direction of the second outlet channel 12. In practice, the first laser pulse 41 creates a first zone of increased viscosity before the entrance to the first outlet channel 11. The sample liquid 20 is thus diverted towards the second outlet channel 12. The first pulse begins to divert the flow from the first outlet channel 11 (waste arm) towards the second outlet channel 12 (sample arm) which sees its flow rate increase. Shortly or immediately after, the second laser pulse 42 has the effect of compensating for the initial deflection and redirecting the sample liquid towards the first outlet channel 11 while retaining only a very small portion 130 of sample liquid in the second outlet channel 12 (sample arm). Indeed, the flow towards the first outlet channel 11 is then restored by a strong decrease in viscosity in the first sheath liquid 21 which causes a return of the flow in the direction of the first outlet channel 11 for the duration of the second pulse.This sequence of two successive laser pulses results in the cutting of a small quantity or portion 130 of sample liquid which is directed towards the second outlet channel 12 (collection or collection arm) and the reestablishment of the main flow of sample liquid towards the first outlet channel 11 or waste arm. This device and this method thus make it possible to extract a volume of sample liquid from a few femtoliters to a few hundred femtoliters in a very short time, because it takes place without interrupting the overall flow in each of the arms. Reducing the duration of the pulses by increasing the power of the laser pulses at constant energy makes it possible to further reduce the duration of the extraction of the portion of sample fluid and thus to further reduce the volume collected.

[0104] This sequence of two successive laser pulses works well because the local temperature variation is very low, of the order of 20°C, which allows low inertia and very fast dynamic behavior of the microfluidic manipulation process. In the example above, between [Fig.3] and [Fig.6], 150ps elapsed. It should also be noted that the sudden increase in viscosity within a very fluid liquid acts as a kind of deflector, which is very different from what could happen if the viscosity decreased. The device and method thus have better efficiency than prior art microfluidic manipulation devices and methods based on a local decrease in viscosity.

[0105] The two-pulse method is applied in a similar manner with heating zones 31, 32 each comprising an electro-thermal transducer, by applying electrical pulses instead of laser pulses. The electrical pulses may have a duration in the millisecond range, less than or equal to 10 ms, 1 ms, 100 s to 10 s.

[0106] In the example of [Fig.l], the two heating zones 31, 32 are arranged symmetrically with respect to the longitudinal axis 14 of the common channel 4 and at the same distance P1=P2= approximately 5 pm from the tip of Injunction Y.

[0107] According to a variant, the heating zones 31, 32 are offset from each other relative to the other along the longitudinal axis 14 of the common channel 4. For example, the first heating zone 31 is closer to the Y junction than the second heating zone 32. The first laser pulse 41 is successively applied in the first heating zone 31 for a duration of 50ps and then, after an interval of 50ps, the second laser pulse 42 in the second heating zone 32 for a duration of 50ps. The first heating zone 31 is for example located just before the inlet of the first outlet channel 11. The second heating zone 32 is located upstream of the first heating zone 31 along the longitudinal axis 14. For example, the first heating zone 31 is at a distance PI = 2pm from the tip of the Y junction and the second heating zone 32 is at a distance P2 = 7pm from the tip of the Y junction.The first laser pulse 41a has the effect of locally and temporarily increasing the viscosity of a portion 111 of the sheath liquid 21 at the inlet of the first outlet channel 11 and of diverting the flow of the sample liquid towards the second outlet channel 12. After 50ps, the second laser pulse 42 has the effect of locally and temporarily increasing the viscosity of another portion 112 of the sheath liquid 22 in the common channel which restores the flow of the entire stream of sample liquid towards the waste channel 11.

[0108] According to a second embodiment illustrated in Figures 7 and 8, a single laser pulse focused on a single heating zone 31 is used. In this embodiment, the sheath liquid 21 comprises a thermoviscous liquid. On the other hand, the sheath liquid 22 does not necessarily comprise a thermoviscous liquid. However, it remains advantageous to use the same sheath liquid 22 as the sheath liquid 21 based on thermoviscous liquid.

[0109] The operating parameters of the laser (power, energy per pulse, pulse duration, wavelength, etc.) are identical to those described in connection with the first pulse of the first embodiment.

[0110] Figures 7 and 8 show the flow of the fluid vein of the sample liquid as well as the temperature of the sheath liquid (the temperature rise being represented by the intensity of the black color in the sheath liquid 21).

[0111] [Fig.7] shows a simulation image of the 50ps microfluidic device after applying a laser pulse 41 to the heating zone 31 for a duration of 50ps. The laser pulse 41 has the effect of locally increasing the temperature to a high temperature of approximately 40°C and thus increasing the viscosity of the sheath liquid 21. The laser pulse 41 creates a zone of increased viscosity which locally and briefly disrupts the flow of the sheath liquid 21 and the sample liquid 20 upstream of the branch between the two outlet channels. In particular, the zone of increased viscosity reduces the flow rate of the sheath liquid 21 to the first outlet channel 11 and diverts the flow of the sample liquid to the second outlet channel 12. It is also observed in [Fig.7] that the temperature rise induced by the laser pulse remains localized in a zone of approximately 10 pm in diameter in the sheath liquid 21. The high temperature reaches approximately 40°C in the heating zone of the sheath liquid 21. On the other hand, the sample liquid undergoes practically no temperature rise, in particular around the heating zone 31.

[0112] Stopping the single laser pulse 41 has the effect of cutting out a small quantity of sample liquid which is directed towards the collection arm ([Fig.8], approximately 100 fps after [Fig.7]). It can be seen in [Fig.8] that the sample liquid is discontinuous in the first outlet channel 11. This discontinuity of the fluid stream in the first outlet channel 11 corresponds to the extraction of the portion 130 of sample liquid towards the second outlet channel 12. The sample liquid therefore flows uninterruptedly. It can also be seen in [Fig.8] that the temperature of the sheath liquid 21 has returned practically to ambient temperature. In addition, the portion 130 of sample liquid extracted in the second outlet channel 12 does not undergo any temperature rise, likely to damage the sample liquid and / or the sorted particles. Likewise, the sample liquid 20 upstream of the heating zone 31 does not undergo any temperature increase.

[0113] In the second embodiment, the restoration of the flow after the heating laser pulse is passive and slower than in the two-pulse configuration. The second embodiment is simpler because it uses a single laser pulse in a single heating zone, but less rapid and less precise than the first embodiment. Indeed, in the first embodiment which uses two pulses, the second pulse is used to force the restoration of the initial flow. The first embodiment is therefore faster and also more precise, since the second pulse allows cutting of the sample vein which has passed into the collection arm. In the second embodiment, this cutting is progressive as the flow is restored.

[0114] In all embodiments, the short distance between the detection unit 50 and the heating zone(s) 31, 32 makes it possible to apply a laser beam very soon after the detection of a particle of interest, which makes it possible to reduce the switching time for extracting the detected particle to the collection channel. In other words, this configuration makes it possible to increase the frequency of switching or sorting or particle extraction.

[0115] The present disclosure advantageously makes it possible to collect single particles from a concentrated liquid.

[0116] In addition, heating the thermo-viscous liquid in the common channel, before entering the outlet channels, makes it possible to obtain significant performance in terms of sorting rate and smallness of the volume collected. The method allows a high flow speed, due to the low viscosity at the injection temperature. The flow speed is in fact of the order of several tens of cm per second, which is very high given the geometry and small dimensions of the microfluidic device. The transient increase in viscosity localized in the sheath liquid is obtained with a low temperature rise (less than 40°C, preferably less than 30°C, or 20°C, or even less than 10°C during switching), which avoids harmful heating of the sample liquid. The method makes it possible to obtain a very high switching speed, due to the small temperature range of viscosity variation, which allows the temperature to be increased and decreased quickly without having to dissipate large amounts of energy, and which limits the thermal inertia of the device and the method.

[0117] The laser requires low optical power (of the order of a few mW) to perform the switching.

[0118] The device and method of the present disclosure make it possible to extract a very small volume (of the order of a few tens of femtoliters), while maintaining a high flow rate of the liquid necessary for analyzing large volumes.

[0119] The use of a thermo-viscous sheath liquid and heating zones upstream of the branch between the outlet channels allows rapid switching without interruption of the sample liquid flow.

Claims

Claims

1. A microfluidic manipulation method comprising the following steps: (a) injecting via an inlet channel (3) a sample liquid (20) into a common channel (4) of a microfluidic device; (b) injecting via at least one other inlet channel (1, 2) at least one sheath liquid (21, 22) at a determined injection temperature into the common channel (4) to allow hydrodynamic focusing of the sample liquid (20) in the common channel (4) and flow of the sample liquid (20) towards a first outlet channel (11) which is located downstream of a branch between the first outlet channel (11) and a second outlet channel (12), the at least one sheath liquid comprising a thermo-viscous liquid, said at least one sheath liquid (21, 22) comprising the thermo-viscous liquid having a dynamic viscosity of less than 0.1 Pa.s at the injection temperature; characterized in that the method comprises the following step: (c) applying an energy source for a limited duration, less than or equal to 10 ms, in a heating zone (31) located in the common channel (4) upstream of the branch between the first outlet channel (11) and the second outlet channel (12), said heating zone (31) being located in or in contact with the at least one sheath liquid (21, 22) comprising the thermo-viscous liquid so as to locally heat said at least one sheath liquid (21, 22) to a high temperature Th in the heating zone (31) of the common channel (4), the injection temperature and the high temperature being separated by a temperature difference less than or equal to 40°C, said at least one sheath liquid comprising the thermo-viscous liquid having a dynamic viscosity greater than or equal to 1.0 Pa.s at the high temperature Th, so as to divert or extract a portion (120, 130, 220) of the sample liquid selectively to the second output channel (12).

2. A method according to claim 1 comprising a following step after step (c): (d) applying another energy source for a limited duration, less than or equal to 10 ms, in a second heating zone (32) located in the common channel (4) upstream of the branch between the first outlet channel (11) and the second outlet channel (12), the second heating zone (32) being located in or in contact with the at least one sheath liquid (21, 22) comprising the thermoviscous liquid for locally heating said at least one sheath liquid (21, 22) to the high temperature Th in the second heating zone (32) of the common channel (4) so ​​as to restore the flow of the sample liquid towards the first outlet channel (11).

3. Method according to claim 2 in which step d) is separated from step c) by a duration less than or equal to 500 ps.

4. Method according to one of claims 1 to 3 in which the energy source of step c) comprises a laser emitting a first laser pulse at a wavelength between 100 nm and 10 pm.

5. The method of claim 2 wherein the further energy source of step d) comprises a laser emitting a second laser pulse at a wavelength between 100 nm and 10 pm.

6. Method according to one of claims 4 to 5 in which the sheath liquid comprising the thermo-viscous liquid is capable of absorbing the first laser pulse and / or the second laser pulse.

7. A method according to claim 4 or, respectively, claim 5, wherein the heating zone (31) or, respectively, the second heating zone (32) comprises a photo-thermal transducer.

8. Method according to one of claims 1 to 3 in which the heating zone (31) or, respectively, the second heating zone (32), comprises an electro-thermal transducer and in which the energy source of step c), respectively of step d), comprises an electrical energy source.

9. Method according to one of claims 1 to 8 in which the thermal variation of dynamic viscosity of the sheath liquid comprising the thermo-viscous liquid has a maximum greater than or equal to 3 Pa.sK 1 in a determined temperature range extending from the injection temperature, over approximately 10 degrees, or 15 degrees, 20 degrees, 25 degrees or even 30 degrees, 35 degrees or 40 degrees above the injection temperature.

10. Method according to one of claims 1 to 9 in which the sample liquid (20) is free of thermo-viscous liquid.

11. Microfluidic device (100) comprising an inlet channel (3) for sample liquid, at least one other inlet channel (1, 2) for sheath liquid, a first outlet channel (11), a second outlet channel (12), a common channel (4) arranged between said inlet channels (1, 2, 3) and said outlet channels (11, 12), the common channel (4) being connected fin- specifically to said inlet and outlet channels (1, 2, 3, 11, 12), a branch separating downstream the first outlet channel (11) from the second outlet channel (12), the sample liquid inlet channel (3) containing a sample liquid (20) injected into the common channel (4), said at least one other inlet channel (1, 2) containing at least one sheath liquid (21, 22) injected into the common channel (4) at a determined injection temperature, the at least one sheath liquid comprising a thermo-viscous liquid, said at least one sheath liquid (21, 22) comprising the thermo-viscous liquid having a dynamic viscosity of less than 0.1 Pa.s at the injection temperature, so as to allow hydrodynamic focusing of the sample liquid (20) in the common channel (4), the microfluidic device being configured to conduct the sample liquid (20) hydrodynamically focused in the common channel (4) by the at least one sheath liquid (21, 22) at the injection temperature towards the first outlet channel (H), characterized in that:. the microfluidic device comprises heating means comprising an energy source (40) and at least one heating zone (31) located in the common channel (4) upstream of the branch between the first outlet channel (11) and the second outlet channel (12), said at least one heating zone (31) being located in or in contact with the at least one sheath liquid (21, 22) comprising the thermo-viscous liquid, the heating means being configured to apply the energy source for a limited duration, less than or equal to 10 ms, in said at least one heating zone (31), so as to locally heat said at least one sheath liquid (21, 22) to a high temperature Th in the common channel (4) in said at least one heating zone (31) of the common channel (4), the injection temperature and the high temperature being separated by a temperature difference less than or equal to 40°C,and in that said at least one sheath liquid comprising the thermo-viscous liquid has a dynamic viscosity greater than or equal to 1.0 Pa.s at the high temperature Th, so as to deflect or extract a portion (120, 130, 220) of the sample liquid selectively towards the second outlet channel (12).,

12. A microfluidic device according to claim 11 wherein the energy source comprises a pulsed laser capable of generating pulses having an energy per pulse between lOnJ and lOOOOnJ.