Microdroplet generation method and generation system

Abstract

This application presents a micro-droplet generating method and a micro-droplet generating system, which can generate a large amount of micro-droplets 201 in a short time, greatly shorten the droplet generating time, and the operation process is simple and convenient. Since equipment such as a high-precision micropump is not required, the system cost can be reduced. In addition, the method is scalable, and more micro-droplets and multiple sets of samples can be separated by expanding the size of the microfluidic chip. In addition, the volume and density of the formed micro-droplets can be precisely adjusted by adjusting the gap between the upper plate and the lower plate, the number of suction points, the area and the position manner. Therefore, the present invention presents a micro-droplet generating method and a micro-droplet generating system that can rapidly form high-density micro-droplets and precisely control the volume and density of the formed high-density micro-droplets.

Description

[Technical field]

[0001] The present invention relates to the field of droplet control technology, and in particular to a method and system for generating micro-droplets. [Background technology]

[0002] How to break down a certain volume of liquid into a large amount of uniformly sized microdroplets is a key problem to be solved in microfluidic control technology, and is an important part of application fields such as droplet digital PCR (dd PCR), dLAMP, dELISA, and single cell genomics. Currently, the main technical means for generating normal liter droplets at high throughput are microdroplet microfluidic control technology and microwell microfluidic control technology. Representative microfluidic control technologies are Bio-Rad and 10XGenomics, and the characteristic of this technology is that it controls a high-precision micropump and uses a cross-shaped structure to continuously press and squeeze the sample liquid to generate a large amount of small droplets ranging from petaliters to normal liters. The method of generating normal liter droplets at high throughput based on microdroplet microfluidic control technology relies on precise control of the pressure of a high-precision micropump and high-precision MEMS chip processing method, and the generated microdroplets are stored together in one container, and the droplets are passed through a microfluidic passage one by one and are inspected one by one, which results in high equipment costs and a complex system. Thermo Fisher is a representative of microwell-based microfluidic control, which uses mechanical force to apply sample liquid to a microwell array, so that the sample is evenly distributed into each microwell, forming small droplets of petaliter to normal liter size. Technologies based on microwell-based microfluidic control usually use mechanical force to apply reagents to the surface of the microwell array, and then fill the upper and lower sides of the microwells with inert liquid. This method has the disadvantages of a relatively complicated operation flow, a low level of automation, a relatively low experimental flow rate, and a long sample preparation period.

[0003] Digital microfluidic control is another technology for generating microdroplets with high throughput because it has the ability to independently control each droplet. Patents WO2016 / 170109Al and US20200061620S50 both describe a method for generating a large amount of microdroplets based on a digital microfluidic control platform. However, the method for generating normal liter droplets with high throughput using digital microfluidic control technology described in the above patents mainly uses digital microfluidic control technology to generate small droplets from large droplets and control them to be transported to the corresponding positions. This method mainly has the disadvantages of a relatively slow speed for generating small droplets and a long sample preparation period. Summary of the Invention [Problem to be solved by the invention]

[0004] In light of the above, it is necessary to develop a method and system for generating microdroplets that can generate small droplets quickly, stably, and controllably.

[0005] The micro-droplet generation system has the following features: it includes a micro-fluidic chip and a droplet driving unit connected to the micro-fluidic chip, the micro-fluidic chip includes an upper plate and a lower plate, a fluid passage layer is formed between the upper plate and the lower plate, the suction point is used to suck liquid, and the droplet driving unit is used to drive the flow of liquid poured into the fluid passage layer in the fluid passage layer, thereby forming micro-droplets at the position of the suction point.

[0006] In one embodiment of the present application, the upper plate comprises an upper cover, a conductive layer and a first hydrophobic layer, which are stacked in order; the lower plate comprises a second hydrophobic layer, a dielectric layer, an electrode layer and a substrate, which are stacked in order; the first hydrophobic layer and the second hydrophobic layer are arranged opposite each other; the fluid passage layer is formed between the first hydrophobic layer and the second hydrophobic layer; and the electrode layer comprises a plurality of electrodes arranged in an array.

[0007] In one embodiment of the present application, the upper plate has a hydrophilic point array formed on the side of the first hydrophobic layer away from the conductive layer, and the hydrophilic points of the hydrophilic point array are the attraction points, with adjacent turned-on electrodes separated by turned-off electrodes.

[0008] In one embodiment of the present application, the upper plate has a hydrophilic point array formed on the side of the first hydrophobic layer away from the conductive layer, and the hydrophilic points of the hydrophilic point array are the attraction points, and the attraction points are formed from the electrodes that are turned on in the electrode layer, with adjacent turned-on electrodes separated by electrodes that are not turned on.

[0009] In one embodiment of the present application, the shape of the electrodes of the electrode layer is hexagonal and square, or hexagonal or square.

[0010] In one embodiment of the present application, the electrode layer consists of a plurality of hexagonal electrodes arranged in an array.

[0011] In one embodiment of the present application, the electrode layer includes a plurality of hexagonal electrodes arranged in an array and a plurality of square electrodes arranged in an array on both sides of the plurality of hexagonal electrodes arranged in an array.

[0012] In one embodiment of the present application, the electrode layer includes a plurality of square electrodes arranged in an array and a plurality of hexagonal electrodes arranged in an array on either side of the plurality of square electrodes arranged in an array.

[0013] In one embodiment of the present application, the side length of the hexagonal electrode is 50 μm to 2 mm, and the side length of the square electrode is 50 μm to 2 mm.

[0014] In one embodiment of the present application, the electrode layer includes a plurality of first square electrodes arranged in an array, a plurality of first hexagonal electrodes arranged in an array, a plurality of second hexagonal electrodes arranged in an array, and a plurality of second square electrodes arranged in an array, which are connected in sequence.

[0015] In one embodiment of the present application, the electrode layer includes a plurality of first hexagonal electrodes arranged in an array, a plurality of second hexagonal electrodes arranged in an array, and a plurality of square electrodes arranged in an array, which are connected in sequence.

[0016] In one embodiment of the present application, the first square electrode or the side length of the square electrode is 50 μm to 2 mm, the side length of the second square electrode is 1 / 5 to 1 / 2 of the side length of the first square electrode, the side length of the first hexagonal electrode is 50 μm to 2 mm, and the side length of the second hexagonal electrode is 1 / 5 to 1 / 2 of the side length of the first hexagon.

[0017] In one embodiment of the present application, the droplet driving unit is an electrode driving unit, which is connected to the electrode layer and is used to control the on and off of the electrodes of the electrode layer, thereby controlling the flow of the liquid injected into the fluid passage layer in the fluid passage layer to form micro-droplets at the suction point.

[0018] In one embodiment of the present application, a liquid discharge hole is provided at the center of the microfluidic chip, and the liquid injection hole is used to inject liquid into the fluid passage layer; the microfluidic chip is provided with a plurality of liquid discharge holes, and the liquid discharge holes are used to discharge excess liquid from the microfluidic chip; the droplet driving unit is a rotation driving unit, and the rotation driving unit is used to drive the rotation of the microfluidic chip.

[0019] In one embodiment of the present application, the rotation speed of the microfluidic chip is greater than 0 rpm and equal to or less than 1000 rpm, so that the liquid injected into the fluid passage layer forms micro-droplets at the suction points by a spin coating method.

[0020] In one embodiment of the present application, the electrode has a hexagonal shape, with a side length of 50 μm to 2 mm, and the distance between the first hydrophobic layer and the second hydrophobic layer is 50 μm to 2 mm.

[0021] In one embodiment of the present application, the microfluidic chip has a first sample inlet hole and a first sample outlet hole, the first sample inlet hole and the first sample outlet hole are arranged on a first diagonal of the microfluidic chip, the droplet driving unit has a first micropump and a third micropump, the first micropump is connected to the first sample inlet hole and is used to inject liquid into the fluid passage layer, thereby filling the fluid passage layer with the liquid, and the third micropump is connected to the first sample outlet hole and draws up the liquid or gas flowing out from the first sample outlet hole, thereby forming a microdroplet at the suction point.

[0022] In one embodiment of the present application, the microfluidic chip has a second sample inlet hole and a second sample outlet hole, the second sample inlet hole and the second sample outlet hole are arranged on a second diagonal of the microfluidic chip, the droplet driving unit has a second micropump and a fourth micropump, the second micropump is connected to the second sample inlet hole and is used to inject liquid into the fluid passage layer, thereby filling the fluid passage layer with the liquid, and the fourth micropump is connected to the second sample outlet hole and draws up the liquid or gas flowing out from the second sample outlet hole, thereby forming a microdroplet at the suction point.

[0023] In one embodiment of the present application, the thickness of the cover thereon is 0.05mm-1.7mm, the thickness of the substrate is 0.05mm-1.7mm, the thickness of the conductive layer is 10nm-500nm, the thickness of the dielectric layer is 50nm-1000nm, the thickness of the electrode layer is 10nm-1000nm, the thickness of the first hydrophobic layer is 10nm-200nm, and the thickness of the second hydrophobic layer is 10nm-200nm.

[0024] The micro-droplet generating system has the following features: A micro-fluidic chip is provided, which is made up of an upper plate and a lower plate, between which a fluid passage layer is formed, and at least one of the upper plate and the lower plate forms a plurality of suction points, which are used to suction liquid. An angle is set between a plane on which the upper plate 10 is located and a plane on which the lower plate 20 is located. The upper plate is provided with a plurality of sample injection holes, which are located at the ends of the upper plate, and which are used to inject liquid. The fluid passage layer includes a first end and a second end, which are disposed opposite to each other. The height of the first end of the fluid passage layer is lower than the height of the second end of the fluid passage layer, and when liquid is injected into the first end of the fluid passage layer through the sample injection holes, the liquid is subjected to surface tension and moves from the first end to the second end, forming micro-droplets at the suction points.

[0025] In one embodiment of the present application, the angle between the upper plate 10 and the lower plate 20 is greater than 0° and less than 3°.

[0026] In one embodiment of the present application, at the first end, the distance between the upper electrode plate and the lower electrode plate is 0 μm to 200 μm.

[0027] In one embodiment of the present application, the upper plate comprises an upper cover, a conductive layer and a first hydrophobic layer, which are stacked in order; the lower plate comprises a second hydrophobic layer, a dielectric layer, an electrode layer and a substrate, which are stacked in order; the first hydrophobic layer and the second hydrophobic layer are arranged opposite each other; the fluid passage layer is formed between the first hydrophobic layer and the second hydrophobic layer; and the electrode layer comprises a plurality of electrodes arranged in an array.

[0028] In one embodiment of the present application, the attraction point is formed from the electrodes of the electrode layer that are turned on, with adjacent turned-on electrodes separated by turned-off electrodes.

[0029] In one embodiment of the present application, the upper plate has a hydrophilic point array formed on the side of the first hydrophobic layer away from the conductive layer, and the hydrophilic points of the hydrophilic point array are the attraction points, and the attraction points are formed from the electrodes that are turned on in the electrode layer, with adjacent turned-on electrodes separated by electrodes that are not turned on.

[0030] In one embodiment of the present application, the shape of the electrodes of the electrode layer is hexagonal and square, or hexagonal or square.

[0031] The microdroplet generation method includes the following steps: Step S1: providing a microfluidic chip. The microfluidic chip includes an upper plate and a lower plate, and a fluid passage layer is formed between the upper plate and the lower plate. Step S2: at least one of the upper plate and the lower plate forms a plurality of suction points, which are used to suck liquid. Step S3: Inject liquid into the fluid passage layer. Step S4: Liquid is driven to flow in the fluid passage layer to form micro-droplets at multiple suction points of the microfluidic chip.

[0032] In one embodiment of the present application, the upper plate comprises an upper cover, a conductive layer and a first hydrophobic layer, which are stacked in sequence; the lower plate comprises a second hydrophobic layer, a dielectric layer, an electrode layer and a substrate, which are stacked in sequence; the electrode layer comprises a plurality of electrodes arranged in an array; and the fluid passage layer is formed between the first hydrophobic layer and the second hydrophobic layer.

[0033] Step S2 includes the steps of turning on a plurality of electrodes of the electrode layer, the turned-on electrodes forming the attraction points and separating adjacent turned-on electrodes by electrodes that are not turned on.

[0034] In one embodiment of the present application, the upper plate comprises an upper cover, a conductive layer and a first hydrophobic layer, which are stacked in sequence; the lower plate comprises a second hydrophobic layer, a dielectric layer, an electrode layer and a substrate, which are stacked in sequence; the electrode layer comprises a plurality of electrodes arranged in an array; and the fluid passage layer is formed between the first hydrophobic layer and the second hydrophobic layer.

[0035] The step S2 includes the following steps: using laser or plasma to process hydrophobic coating at required positions, to form hydrophilic points in the first hydrophobic layer, which are the suction points, and adjacent hydrophilic points are spaced apart from each other.

[0036] In one embodiment of the present application, the above step S4 includes the following steps:

[0037] In step S110, the first to Pth electrodes are turned on, so that the liquid forms large droplets at positions corresponding to the first to Pth electrodes of the fluid passage layer, where P is a positive integer. In step S120, the electrodes of the attraction points in the first row are kept on, and the other electrodes in the first row are turned off, while the electrodes in the P+1 row are turned on to drive the large droplets to move forward one row in the fluid passage layer, and the attraction points in the first row form micro droplets, separated from each other by at least one electrode. In step S130, the electrodes of the attraction points in the second row are kept on, and the other electrodes in the second row are turned off, while the electrodes in the P+2 row are turned on to drive the large droplets to move forward one row in the fluid passage layer; the attraction points in the second row form micro droplets, and there is at least one electrode separating each of the adjacent attraction points; and the attraction points in the first row and the attraction points in the second row are in different rows. In step S140, the electrode of the attraction point in the nth column is kept on, and the other electrodes in the nth column are turned off, while the electrode in the P+nth column is turned on to drive the large droplet to move forward one column in the fluid passage layer, and the attraction point in the nth column forms a micro droplet, and is separated from the adjacent attraction point by at least one electrode, the attraction point in the nth column and the attraction point in the n-1th column are in different columns, and n is a positive integer greater than 3. Steps S150 and S140 are repeated to form multiple micro-droplets in the microfluidic chip until the large droplets are exhausted.

[0038] In one embodiment of the present application, the above step S4 includes the following steps:

[0039] In step S210, the electrodes in the first to Pth rows are turned on, so that the liquid in the fluid passage layer 101 forms large droplets on the electrodes in the first to Pth rows of the electrode layer, where P is a positive integer. In step S220, the electrodes in the first row are turned off and the electrodes in the P+1 row are turned on at the same time, driving the large droplets to move forward one row in the fluid passage layer, thereby forming micro-droplets at the hydrophilic points in the first row. In step S230, the electrodes in the first row are turned off and the electrodes in the P+2 row are turned on at the same time, driving the large droplets to move forward one row on the electrode layer, and forming micro-droplets at the hydrophilic points in the second row. In step S240, the nth electrode is turned off and the P+nth electrode is turned on at the same time to drive the large droplet to move forward one row on the electrode layer, thereby forming a micro-droplet at the nth hydrophilic point, where n is a positive integer greater than 3. Steps S250 and S240 are repeated to form multiple micro-droplets in the microfluidic chip until the large droplets are exhausted.

[0040] In one embodiment of the present application, step S4 includes the steps of: rotating the microfluidic chip to form micro-droplets at positions corresponding to a plurality of liquid-on electrodes in the fluid passage layer.

[0041] In one embodiment of the present application, the step S4 includes the steps of: rotating the microfluidic chip so that the liquid in the fluid passage layer forms micro-droplets at the positions of the hydrophilic points.

[0042] In one embodiment of the present application, in step S4, the rotation speed of the microfluidic chip is greater than 0 rpm and less than or equal to 1000 rpm.

[0043] In one embodiment of the present application, in step S3, liquid is injected into a liquid injection hole located at the center of the microfluidic chip.

[0044] In one embodiment of the present application, the micro-droplet generating method further includes the following step: stopping rotating the micro-fluidic chip after excess liquid has flowed out of the fluid passage layer.

[0045] In one embodiment of the present application, an angle is set between the plane on which the upper plate is located and the plane on which the lower plate is located; a plurality of sample injection holes are provided in the upper plate, the sample injection holes are located at the ends of the upper plate, and the sample injection holes are used for injecting samples; the fluid passage layer includes a first end and a second end arranged opposite to each other, and the height of the first end of the fluid passage layer is lower than the height of the second end of the fluid passage layer.

[0046] In one embodiment of the present application, liquid is injected from the sample injection hole into the first end of the fluid passage layer, and when the liquid is injected into the fluid passage layer, the liquid is subjected to surface tension and moves from the first end to the second end, forming a microdroplet at a position corresponding to the suction point.

[0047] In one embodiment of the present application, in step S3, a liquid is injected from the sample injection hole into the first end of the fluid passage layer, and the injection speed of the liquid is 1 μL / s to 10 μL / s.

[0048] In one embodiment of the present application, at the first end, the distance between the upper electrode plate 10 and the lower electrode plate 20 is 0 μm to 200 μm. The angle between the upper electrode plate and the lower electrode plate is greater than 0° and less than 3°.

[0049] In one embodiment of the present application, the microfluidic chip has a first sample inlet hole and a first sample outlet hole, the first sample outlet hole and the first sample inlet hole are arranged on a first diagonal of the microfluidic chip, the first sample inlet hole is connected to a first micropump, and the first sample outlet hole is connected to a third micropump.

[0050] In step S3, a liquid is injected from a first micropump into the fluid passage layer via a first sample injection hole, and the liquid discharged from the first sample discharge hole is extracted by a third micropump.

[0051] In one embodiment of the present application, the microfluidic chip has a second sample inlet hole and a second sample outlet hole, the second sample outlet hole and the second sample inlet hole are arranged on a second diagonal of the microfluidic chip, the second sample inlet hole is connected to a second micropump, and the second sample outlet hole is connected to a fourth micropump.

[0052] In step S4, a medium is injected from a second micropump into the fluid passage layer via a second sample injection hole, the liquid at the non-suction point is pushed into the medium, the liquid leaves a micro-droplet at the position corresponding to the suction point, and the medium envelops the micro-droplet, and then a fourth micropump pumps up the medium from the second sample discharge hole.

[0053] In one embodiment of the present application, the volume and density of micro-droplets formed by the microfluidic chip are adjusted by adjusting the gap between the upper plate and the lower plate, the number, area size and position of the suction points.

[0054] A method for generating microdroplets, comprising the steps of: A microfluidic chip is provided, the microfluidic chip including an upper plate and a lower plate, between which a fluid passage layer is formed, the lower plate including an electrode layer, the electrode layer including a plurality of electrodes arranged in an array.

[0055] The upper plate forms a plurality of attraction points, which are used to attract liquid, and the attraction points are formed by electrodes of the electrode layer that are turned on, and adjacent electrodes that are turned on are spaced apart by electrodes that are not turned on.

[0056] A liquid sample is injected into the fluid passage layer, and the electrodes are controlled to be turned on and off, so that the liquid sample forms n1 micro-droplets at the positions corresponding to the attraction points.

[0057] In addition, by controlling the on and off of the electrodes, all of the n1 microdroplets formed form n2 microdroplets at the attraction point.

[0058] Furthermore, by controlling the on and off of the electrodes, all of the n2 microdroplets formed form n3 microdroplets at the attraction point.

[0059] The electrodes are cycled on and off until the desired number of microdroplets is formed.

[0060] Among them, n1, n2, and n3 are positive integers equal to or greater than 2.

[0061] In one embodiment of the present application, a liquid sample is injected into the fluid passage layer, and by controlling the electrodes to be on and off, the liquid sample forms two micro-droplets at positions corresponding to the attraction points.

[0062] Furthermore, by controlling the on and off of the electrodes, all of the two microdroplets formed form two microdroplets at the attraction point.

[0063] Furthermore, by controlling the on and off of the electrodes, all of the two microdroplets formed form two microdroplets at the attraction point.

[0064] The electrodes are cycled on and off until the desired number of microdroplets is formed.

[0065] In one embodiment of the present application, a liquid sample is injected into the fluid passage layer, and by controlling the electrodes to be on and off, the liquid sample forms three micro-droplets at positions corresponding to the attraction points.

[0066] In addition, by controlling the on and off of the electrodes, all of the three microdroplets formed form three microdroplets at the attraction point.

[0067] Furthermore, by controlling the on and off of the electrodes, all of the three formed microdroplets form three microdroplets at the attraction point.

[0068] The electrodes are cycled on and off until the desired number of microdroplets is formed.

[0069] In one embodiment of the present application, a liquid sample is injected into the fluid passage layer, and by controlling the electrodes to be turned on and off, the liquid sample forms four micro-droplets at positions corresponding to the attraction points.

[0070] In addition, by controlling the on and off of the electrodes, all of the four formed microdroplets form four microdroplets at the attraction point.

[0071] Furthermore, by controlling the on and off of the electrodes, all of the four formed microdroplets form four microdroplets at the attraction point.

[0072] The electrodes are cycled on and off until the desired number of microdroplets is formed.

[0073] In one embodiment of the present application, the electrodes are square and hexagonal in shape.

[0074] In one embodiment of the present application, the upper plate comprises an upper cover, a conductive layer and a first hydrophobic layer, which are stacked in order; the lower plate also includes a second hydrophobic layer and a dielectric layer, in which the second hydrophobic layer, the dielectric layer and the electrode layer are stacked in order; the first hydrophobic layer and the second hydrophobic layer are disposed opposite each other; and the fluid passage layer is formed between the first hydrophobic layer and the second hydrophobic layer.

[0075] In one embodiment of the present application, the side length of the electrode is 50 μm to 2 mm.

[0076] In one embodiment of the present application, the distance between the first hydrophobic layer and the second hydrophobic layer is 5 μm to 600 μm.

[0077] The present application provides a microdroplet generating method and a microdroplet generating system, which can generate a large amount of microdroplets quickly, greatly reduce the droplet generating time, and simplify the operation process. Since no equipment such as a high-precision micropump is required, the system can reduce the cost. The system is also scalable, and can separate more microdroplets and multiple sets of samples by expanding the size of the microfluidic chip. In addition, the volume and density of the formed microdroplets can be precisely adjusted by adjusting the gap between the upper plate and the lower plate, the number of suction points, the area and the position of the suction points. Thus, the present invention provides a microdroplet generating method and a microdroplet generating system, which can rapidly form high-density microdroplets and precisely control the volume and density of the formed high-density microdroplets.

[0078] The microdroplet generating method and generating system of the present application have good scalability, and can separate more microdroplets and multiple sets of samples through the expansion of chip size. In addition, the electrode layer includes at least two different types of electrodes that look like arrays, so that one type of array of electrodes with large droplets can form microdroplets, and another type of array of electrodes can complete the related experiment of microdroplets by turning on and off the electrodes, thus avoiding the transfer of liquid samples. [Brief description of the drawings]

[0079] [Figure 1] 1 is a schematic cross-sectional view of a microfluidic chip of a micro-droplet generating system according to a first embodiment of the present invention. [Diagram 2] 1 is a structural diagram of a micro-droplet generating system according to a first embodiment of the present invention. [Diagram 3] 2 is a flow chart of a micro-generation method using the micro-droplet generation system of FIG. 1. [Figure 4] 1 is a flow chart showing a large droplet moving to form a single microdroplet. [Diagram 5]1 is a flow chart showing a large droplet moving to form multiple microdroplets. [Figure 6] 1 is a flow chart showing a first embodiment of the present invention in which a large droplet moves through a microfluidic chip to form multiple micro-droplets. [Figure 7] 1 is an actual experimental diagram of a first embodiment of the present invention in which a large droplet moves in a microfluidic chip to form multiple micro-droplets. [Figure 8] 1 is a schematic diagram of a large droplet moving in a microfluidic chip to form multiple micro-droplets according to a first embodiment of the present invention. FIG. [Figure 9] 1 is a flowchart of a micro-droplet generating method of a micro-droplet generating system according to a first embodiment of the present invention. [Figure 10] 1 is a diagram illustrating a micro-droplet generating method of a micro-droplet generating system according to a second embodiment of the present invention. [Figure 11-13] 10 is a flowchart of a micro-droplet generating method of a micro-droplet generating system according to a second embodiment of the present invention. [Figure 14] 1 is a structural diagram of a micro-droplet generating system according to a third embodiment of the present invention. [Figure 15] 1 is a schematic cross-sectional view of a microfluidic chip of a micro-droplet generating system according to a third embodiment of the present invention. FIG. [Figure 16-17] 11 is a diagram illustrating a micro-droplet generating method of a micro-droplet generating system according to a third embodiment of the present invention. [Figure 18] This is a structural diagram of the mixed solution in the Elisa. [Figure 19] 1 is a schematic diagram of the operation process of digital Elisa using a micro droplet generation system. [Figure 20-21] 11 is a flowchart of a micro-droplet generating method of a micro-droplet generating system according to a third embodiment of the present invention. [Figure 22-25] 11 is a diagram illustrating a micro-droplet generating method of a micro-droplet generating system according to a fourth embodiment of the present invention. [Figure 26-27]11 is a flowchart of a micro-droplet generating method of a micro-droplet generating system according to a fourth embodiment of the present invention. [Figure 28] 5 is a schematic cross-sectional view of a microfluidic chip of a microdroplet generating system according to a fifth embodiment of the present invention, showing a process of generating microdroplets. FIG. [Figure 29] FIG. 11 is a structural diagram of a first type of electrode layer according to a fifth embodiment of the present invention. [Diagram 30] 1 is a schematic diagram of liquid movement and generation of micro-droplets when the first type of electrode layer according to the fifth embodiment of the present invention is adopted. [Diagram 31] FIG. 11 is a schematic diagram showing a second type of structure of an electrode layer according to a fifth embodiment of the present invention. [Diagram 32] 1 is a schematic diagram of liquid movement and generation of micro-droplets when the second type of electrode layer according to the fifth embodiment of the present invention is adopted. [Diagram 33] FIG. 13 is a schematic diagram showing how liquid moves and generates microdroplets in a fifth embodiment of the present invention, and shows a process for conducting a cell sorting experiment by employing this microdroplet generation method. [Diagram 34] FIG. 5 is a schematic diagram of a fifth embodiment of the present invention in which liquid moves and microdroplets are generated, showing the process of microdroplets forming petaliters. [Diagram 35] 10 is a flowchart of a micro-droplet generating method according to a fifth embodiment of the present invention. [Diagram 36] 11 is a flowchart of a micro-droplet generating method according to a sixth embodiment of the present invention. [Figure 37] FIG. 13 is a schematic diagram of a first type of method for moving a liquid sample and generating micro-droplets in the sixth embodiment of the present invention. [Figure 38] 1 is an experimental diagram of the first method of liquid movement and microdroplet generation in the sixth embodiment of the present invention, illustrating the process of forming petaliter-level microdroplets. [Figure 39] 1 is an experimental diagram of single cell separation using the first method in which liquid moves and microdroplets are generated in the sixth embodiment of the present invention. [Diagram 40]FIG. 13 is a schematic diagram of a second type of method for moving a liquid sample and generating micro-droplets according to the sixth embodiment of the present invention. [Diagram 41] FIG. 13 is a schematic diagram of a third type of method for moving a liquid sample and generating micro-droplets according to the sixth embodiment of the present invention. [Diagram 42] FIG. 13 is a schematic diagram of a fourth type of method for moving a liquid sample and generating micro-droplets according to the sixth embodiment of the present invention. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0080] In order to make the purpose, technical plan and advantages of the present application more comprehensible, the present application will be described in more detail below in conjunction with the figures and embodiments. Please understand that the detailed embodiments described herein are only for illustrating the present application, not for limiting the present application.

[0081] First embodiment As shown in FIG. 1 to FIG. 9, the detailed structure of the micro-droplet generating system and the micro-droplet generating method according to the first embodiment of the present application are interpreted in detail.

[0082] Specifically, the micro-droplet generating system includes a micro-fluidic chip 100 and a droplet driving unit connected to the micro-fluidic chip 100, the micro-fluidic chip 100 includes an upper electrode 10 and a lower electrode 20, a fluid passage layer 101 is formed between the upper electrode 10 and the lower electrode 20, at least one of the upper electrode 10 and the lower electrode 20 forms a plurality of suction points, which are used to absorb liquid 200. The droplet driving unit drives the injected liquid 200 in the fluid passage layer 101 to flow in the fluid passage layer 101, thereby forming micro-droplets 201 at the positions of the suction points.

[0083] More specifically, as shown in Fig. 1, the upper plate 10 includes an upper cover 11, a conductive layer 12 and a first hydrophobic layer 13, which are arranged in sequence. The lower plate 20 includes a second hydrophobic layer 21, a dielectric layer 22 and an electrode layer 23, which are arranged in sequence. The first hydrophobic layer 13 and the second hydrophobic layer 21 are arranged opposite to each other, and a fluid passage layer 101 is formed between the first hydrophobic layer 13 and the second hydrophobic layer 21. At least one of the upper plate 10 and the lower plate 20 forms a plurality of suction points, which are used to attract liquid 200, and the electrode layer 23 includes a plurality of electrodes 24 arranged in an array.

[0084] In this embodiment, the droplet driving unit is the electrode driving unit, and the electrode driving unit is connected to the electrode layer 23. By controlling the on and off of the electrodes 24 of the electrode layer 23, the flow of the liquid 200 injected into the fluid passage layer 101 in the fluid passage layer 101 can be controlled, and a micro-droplet 201 can be formed at the position of the attraction point.

[0085] It should be understood that the specifications of the above-mentioned multiple suction points can be the same or different, and the number and positions can be set according to actual needs, which is convenient for simultaneously generating micro-droplets 201 with the same or different volumes.

[0086] It should also be understood that the volume and density of the micro-droplets 201 formed in the microfluidic chip 100 can be adjusted by controlling the gaps in the fluid passage layer 101 and the number, positions and area of ​​the suction points. Therefore, the present application provides a micro-droplet generating method and a micro-droplet generating system that can quickly form high-density micro-droplets and precisely control the volume and density of the formed high-density micro-droplets.

[0087] Alternatively, as shown in Figures 4 and 5, the attraction points are formed by turned-on electrodes 241 of the electrode layer 23, and adjacent turned-on electrodes 241 are separated by turned-off electrodes 242.

[0088] The shape of the electrodes 24 of the electrode layer 23 can be selected to be hexagonal or square. In this embodiment, the shape of the electrodes 24 is hexagonal. When the shape of the electrodes 24 is hexagonal, the contact surface is larger and the utilization rate of the electrode 24 panel is higher. It should be understood that the shape of the electrodes 24 can be a combination of hexagonal and square, or a combination of any other shape and any other shape. There is no limitation to the above shapes in this application.

[0089] The side length of the hexagonal electrode can be selected from 50 μm to 2 mm, and the side length of the square electrode can be selected from 50 μm to 2 mm. The present application also has no limitation on the size of the electrode 24.

[0090] In the above micro-droplet generating system, large droplets are put into the fluid passage layer 101, and the electrode 24 of the electrode layer 23 is controlled to be turned on and off through the electrode driving unit, so that the large droplets put into the fluid passage layer 101 are controlled to flow on the surface of the electrode layer 23 in a manner similar to coating, and the micro-droplets 201 are formed at multiple suction points of the fluid passage layer 101, which greatly shortens the droplet generation time, improves the droplet generation stability, and can adjust the size of the droplets generated according to demand at any time, and the operation process is simple and convenient. No equipment such as high-precision micro pumps is required, which leads to reduced system costs. In addition, it is highly scalable, and more micro-droplets or multiple sets of samples can be separated by expanding the micro-fluid control size.

[0091] Optionally, as shown in Fig. 2, in one variant of this embodiment, the attraction point can be formed from a hydrophilic point 131. Specifically, the upper electrode 10 has a hydrophilic point array formed on the first hydrophobic layer 13 away from the conductive layer 12, and the hydrophilic point 131 of the hydrophilic point array is the attraction point and is located between the adjacent hydrophilic points 131.

[0092] It should be understood that the hydrophilic dot array can also be formed in the second hydrophobic layer 21. Or the first hydrophobic layer 13 and the second hydrophobic layer 21 are both provided with hydrophilic dots 131. This application is not limited to this item.

[0093] 2, through hydrophilic modification, the first hydrophobic layer 13 forms a hydrophilic dot array on the side away from the conductive layer 12, and at least one electrode 24 is spaced between adjacent hydrophilic dots 131. An electrode driving unit is connected to the electrode layer 23, and the electrode driving unit is used to move the large droplets into the fluid passage layer 101. The large droplets form microdroplets 201 at the hydrophilic dots 131. It should be understood that the volume of the microdroplets 201 formed by the microdroplet generating system depends on the size of the gap h of the fluid passage layer 101 and the area of ​​the hydrophilic dots 131.

[0094] In the above micro-droplet generating system, the electrode driving unit is used to move the large droplets in the fluid passage layer 101 through putting the large droplets into the fluid passage layer 101, and when the large droplets pass through the hydrophilic point 131, the micro-droplets 201 are left at the hydrophilic point 131 due to the hydrophilicity of the hydrophilic point 131. This greatly shortens the droplet generation time. In addition, the above micro-droplet generating system does not need to control the electrodes to separate the micro-droplets 201, making the operation more convenient. No equipment such as a high-precision micropump is required, which leads to a reduction in system costs. In addition, the system is highly scalable, and more micro-droplets or multiple sets of samples can be separated by expanding the microfluidic control size.

[0095] It should be understood that the present application also provides a method for generating micro-droplets in the micro-droplet generating system shown in Figure 1, which includes the following steps:

[0096] By controlling the on and off of the electrodes 24 of the electrode layer 23, when the large droplet passes through the electrode layer 23, the micro-droplets 201 are formed at the multiple attraction points of the electrode layer 23 respectively.

[0097] In the above micro-droplet generating method, the electrodes 24 of the electrode layer 23 are turned on and off, and when large droplets flow through the electrode layer 23, micro-droplets 201 are formed at multiple attraction points of the electrode layer 23, respectively, which greatly shortens the droplet generating time and makes the operation process simple and convenient.

[0098] It should be understood that the sizes of the above multiple suction points can be the same or different, which is convenient for generating micro-droplets 201 with different volumes simultaneously.

[0099] In addition, the attraction points are separated by at least one electrode 24. By separating the attraction points by at least one electrode 24, the microdroplets 201 are prevented from combining. More preferably, the attraction points are separated by two electrodes 24.

[0100] The specific operation method is shown in FIG. 3. The electrodes 24 of the electrode layer 23 are controlled to be turned on and off, and when large droplets flow through the electrode layer 23, micro-droplets 201 are formed at multiple attraction points of the electrode layer 23, respectively. The operation is as follows:

[0101] In step S110, the electrodes 24 in the first to Pth rows are turned on, and large droplets of the liquid 200 are formed in the fluid passage layer 101 at positions corresponding to the electrodes 24 in the first to Pth rows, where P is a positive integer.

[0102] In step S120, the electrodes 24 of the attraction points in the first row are kept on, and the other electrodes 24 in the first row are turned off, while the electrodes 24 in the P+1 row are turned on to drive the large droplet to move forward one row in the fluid passage layer 101. In addition, the attraction points in the first row form micro droplets 201, and are separated from each other by at least one electrode 24.

[0103] In step S130, the electrodes 24 of the attraction points in the second row are kept on, and the other electrodes 24 in the second row are turned off, while the electrodes 24 in the P+2 row are turned on to drive the large droplet to move forward one row on the fluid passage layer 101. The attraction points in the second row form micro droplets 201, and are separated from each other by at least one electrode 24. The attraction points in the first row and the attraction points in the second row are in different rows.

[0104] In step S140, the electrode 24 of the attraction point in the nth column is kept on, and the other electrodes 24 in the nth column are turned off, and at the same time, the electrode 24 in the P+nth column is turned on to drive the large droplet to move one column forward in the fluid passage layer 101. In addition, the attraction point in the nth column forms a micro droplet 201, and is separated from the adjacent attraction point by at least one electrode 24. The attraction point in the nth column and the attraction point in the n-1th column are in different columns, and n is a positive integer greater than 3.

[0105] Steps S150 and S140 are repeated to form a plurality of micro-droplets 201 in the microfluidic chip 100 until the large droplets are exhausted.

[0106] It should be understood that the specific operations of repeating step S140 in step S150 are as follows: n=3, execute step S140 once; n=4, execute step S140 once; n=5, execute S140 once. Repeat until the large droplets are used up. That is, the large droplets are moved in order from the first row to the nth row, and multiple micro droplets 201 are formed in each of the first row to the nth row.

[0107] It should be understood that the "discharge" (translated as "row" above) in the above microdroplet generating method can be expressed as "row". That is, large droplets move in order from the first row to the nth row, forming multiple microdroplets 201 from the first row to the nth row.

[0108] In one embodiment, the volume of the microdroplet 201 can be controlled by adjusting the distance between the first hydrophobic layer 13 and the second hydrophobic layer 21 and the size of one electrode 24. By adjusting the distance between the first hydrophobic layer 13 and the second hydrophobic layer 21 and the size of one electrode 24, the volume of the microdroplet 201 can be precisely controlled from the petaliter range to the normal liter range.

[0109] Specifically, referring to Figure 4, an electrode array consisting of electrodes 24 controls the movement of the large droplet along the electrode array in the direction of the arrow in the figure. By controlling the electrode array, a single microdroplet 201 can be separated from the large droplet. The large droplet continues to move in the direction of the arrow, but the microdroplet 201 remains in its original position.

[0110] As shown in Fig. 5, by repeating the operation of Fig. 4, the large droplet leaves several microdroplets 201 on its moving path, and the microdroplets 201 are separated by several electrodes 24 to prevent the microdroplets 201 from joining together. The electrodes 24 located under the microdroplets 201 are turned on to fix the microdroplets 201 at their original positions. It is possible to separate and stop the microdroplets 201 as many as the target number, or to repeat the separation step until the large droplets are completely used up.

[0111] Furthermore, as shown in FIG. 6, the large droplets are controlled in the order of FIG. 6(A) to FIG. 6(F) to leave multiple microdroplets 201 on the passage. The microdroplets 201 are separated by several electrodes 24 to prevent the microdroplets from merging. The electrodes 24 located under the microdroplets 201 are turned on to fix the microdroplets 201 in their original positions. The microdroplets 201 may be separated and stopped as many as the target number of microdroplets 201, or the separation step may be repeated until the large droplets are completely used up. The microdroplets 201 are between the first hydrophobic layer 13 and the second hydrophobic layer 21. By adjusting the gap h of the fluid passage layer 101 and the size of the electrodes 24, the volume of the microdroplets 201 can be precisely controlled from the petaliter level to the normal liter level.

[0112] 7 shows the actual experimental process of a large droplet moving through a microfluidic chip to form multiple microdroplets in the first embodiment of the present invention. The process of a large droplet moving through a microfluidic chip to form multiple microdroplets is consistent with FIG.

[0113] Referring to FIG. 8, when the electrodes 24 are different sizes or when one to multiple adjacent electrodes 24 are turned on, micro-droplets 201 of different sizes can be formed on the electrode layer 23 .

[0114] The present application also provides a method for generating micro-droplets using the micro-droplet generating system shown in Figure 2. The steps are as follows:

[0115] By controlling the electrodes 24 of the electrode layer 23 to be turned on and off, micro-droplets 201 are formed at the hydrophilic dot array of the electrode layer 23 when the larger droplets flow through the electrode layer 23 .

[0116] In one embodiment, the size of the hydrophilic dots 131 is controlled to control the volume of the microdroplets.

[0117] In the above micro-droplet generating method, large droplets are introduced into the fluid passage layer 101, and an electrode driving unit is used to drive the flow of the large droplets in the fluid passage layer 101, and when the large droplets pass through the hydrophilic points 131, the hydrophilic effect of the hydrophilic points 131 causes the micro-droplets 201 to remain at the hydrophilic points 131, thereby greatly shortening the droplet generation time. In addition, the above micro-droplet generating system does not require the electrodes 24 to separate the micro-droplets 201, making the operation more convenient.

[0118] Referring to FIG. 9, by controlling the on and off of the electrodes 24 of the electrode layer 23, when a large droplet flows through the electrode layer 23, the operation of forming a micro-droplet 201 at the hydrophilic point array of the electrode layer 23 is as follows.

[0119] In step S210, the electrodes 24 in the first to Pth rows are turned on, so that the liquid 200 in the fluid passage layer 101 forms large droplets on the electrodes 24 in the first to Pth rows of the electrode layer 23, where P is a positive integer.

[0120] In step S220, the electrodes 24 in the first row are turned off and at the same time, the electrodes 24 in the P+1 row are turned on to drive the large droplet to move forward one row in the fluid passage layer 101, thereby forming micro droplets 201 at the hydrophilic points 131 in the first row.

[0121] In step S230, the electrodes 24 in the first row are turned off and at the same time, the electrodes 24 in the P+2 row are turned on to drive the large droplets to move forward one row on the electrode layer 23, thereby forming micro droplets 201 at the hydrophilic points 131 in the first row.

[0122] In step S240, the n-th electrode 24 is turned off and the P+n-th electrode 24 is turned on at the same time, so that the large droplet is driven to move forward one row on the electrode layer 23, and a micro droplet 201 is formed at the n-th hydrophilic point 131, where n is a positive integer greater than 3.

[0123] S250 and S240 are repeated to form multiple micro-droplets 201 in the microfluidic chip 100 until the large droplets are exhausted.

[0124] It should be understood that the specific operations of repeating S240 in S250 are as follows: n=3, execute S240 once; n=4, execute S240 once; n=5, execute S240 once; ...repeat until the large droplets are used up. That is, the large droplets are moved in order from the first row to the nth row, and multiple micro droplets 201 are formed in each of the first row to the nth row.

[0125] It should be understood that the "discharge" (translated as "row" above) in the above microdroplet generating method can be expressed as "row", that is, the large droplets move in order from the first row to the nth row, and form multiple microdroplets 201 from the first row to the nth row, respectively.

[0126] Using the above-described microdroplet generation method, the separation step can be repeated to separate droplets up to a desired number.

[0127] The above microdroplet generating method is different from the method of generating microdroplets 201 using traditional digital microfluidic control, in which traditional digital microfluidic control steers a large droplet to generate one microdroplet 201 and transports the microdroplet 201 to the destination. The above microdroplet generating method controls the liquid 200 to pass through the fluid passage layer 101, and controls the electrodes 24 to make the large droplet leave the microdroplet 201 on its path. Alternatively, the upper cover 11 is hydrophilically modified in an array manner, and when the large droplet flows through the hydrophilic points 131, the hydrophilic effect of the hydrophilic points 131 causes the microdroplet 201 to remain at the hydrophilic points 131. Compared with the method of generating microdroplets 201 using traditional digital microfluidic control, the above microdroplet generating method can greatly shorten the droplet generation time.

[0128] Through the above-mentioned micro-droplet generation method, large droplets can be controlled to be coated on the electrode layer 23. Through the control of the electrode 24 or the array-type hydrophilic modification of the upper cover 11, high-throughput normal liter-class droplet generation can be realized. The volume of the droplets can be precisely adjusted by adjusting the size of the electrode 24, the gap distance of the electrode 24, or the size of the hydrophilic modification point. After the high-throughput normal liter droplet separation is completed, the corresponding experiment or detection can be performed on the digital microfluidic chip. When this method is used in conjunction with an optical detection module, it can also realize biochemical application functions such as ddPCR, dLAMP, and dELISA single-cell experiments. It can also be applied to other nucleic acid tests, such as isothermal amplification. In addition, the microfluidic chip 100 can be used to perform narrowing down or independent experiments on any micro-droplets, and the size of the microfluidic chip 100 can be expanded to separate more micro-droplets or more groups of samples.

[0129] Second embodiment 10 to 13, the detailed structure of the micro-droplet generating system and the micro-droplet generating method according to the second embodiment of the present application are specifically described. The second embodiment is a modified example of the first embodiment.

[0130] The micro-droplet generating system of the second embodiment includes a microfluidic chip 100 and a droplet driving unit of the microfluidic chip 100. The microfluidic chip 100 includes an upper electrode plate 10 and a lower electrode plate 20, and the lower electrode plate 20 includes a second hydrophobic layer 21, a dielectric layer 22 and an electrode layer 23, which are arranged in sequence. The first hydrophobic layer 13 and the second hydrophobic layer 21 are arranged opposite to each other, the fluid passage layer 101 is formed between the first hydrophobic layer 13 and the second hydrophobic layer 21, and the electrode layer 23 includes a plurality of electrodes 24 arranged in an array. At least one of the upper electrode plate 10 and the lower electrode plate 20 forms a plurality of suction points, which are used to suck the liquid 200. The purpose of the droplet driving unit is to drive the flow of the liquid 200 poured into the fluid passage layer 101 in the fluid passage layer 101. In this way, a micro-droplet 201 is formed at the location of the attraction point.

[0131] 10, a liquid injection hole 132 is provided at the center of the microfluidic chip 100, which is used to inject a liquid 200 into the fluid passage layer 101. The microfluidic chip 100 also has a plurality of liquid discharge holes 133, which are used to discharge the remaining liquid 200 from the microfluidic chip 100. The droplet driving unit is a rotation driving unit, which is used to rotate the microfluidic chip 100. Thus, the liquid 200 injected into the fluid passage layer 101 can be spin-coated to form micro-droplets 201 at the suction points.

[0132] It should be understood that the liquid injection hole 132 is located at the center of the microfluidic chip 100 so that the liquid 20 is evenly injected into the fluid passage layer 101 and the micro-droplets 201 are evenly formed in the microfluidic chip 100 when the microfluidic chip 100 is rotated. In the present application, there are also embodiments in which the liquid injection hole 132 is not located at the center of the microfluidic chip 100. There is no limitation to this in the present application.

[0133] It should be noted that the rotation drive unit can be replaced with a turntable, a rotor, etc., as long as it can rotate the microfluidic chip 100. The present application does not limit the specific structure of the rotation drive unit.

[0134] Specifically, in the order shown in Fig. 10(A) to Fig. 10(F), first, as shown in Fig. 10(A), the microfluidic chip 100 consisting of the electrodes 24 is filled with the liquid 200 to the limit through the liquid injection hole 132. Next, the microfluidic chip 100 rotates in the direction shown by the first arrow 31 in Fig. 10(B), and the liquid 200 starts to move along the microfluidic chip 100 in the direction shown by the second arrow 32 in Fig. 10(B) due to the generation of centrifugal force. The electrodes 24 in the upper part of the microfluidic chip 100 are controlled to be turned on, and as shown in Fig. 10(B), the liquid 200 is left in the next group of microdroplets 201, with the electrodes 242 that are not turned on separating the adjacent turned-on electrodes 241 from each other. As shown in Figures 10(C) to 10(F), the microfluidic chip is continuously rotated, and the liquid 200 continues to be discharged from the liquid discharge holes located at the corners of the array along the direction of the arrows, leaving the microdroplets 201 at the positions of the electrodes 241 that are turned on. By continuously rotating the microfluidic chip 100 and maintaining the centrifugal force, the liquid 200 can leave multiple groups of microdroplets 201 in its discharge passage. The electrodes 24 located under the microdroplets 201 are kept in the turned-on state to fix the microdroplets 201 in their original positions, and the centrifugation continues until the remaining liquid 200 is completely removed and the target number of microdroplets 201 is separated.

[0135] It should be understood that, as shown in FIG. 11, when implementing the second embodiment, the above micro-droplet generating method includes the following steps:

[0136] Step S10: Provide a microfluidic chip 100. The microfluidic chip 100 comprises an upper electrode plate 10 and a lower electrode plate 20. A fluid passage layer is formed between the upper electrode plate 10 and the lower electrode plate 20.

[0137] In step S20, at least one of the upper plate and the lower plate forms a plurality of suction points, which are used to suck the liquid 200.

[0138] In step S30, the liquid 200 is poured into the fluid passage layer 101.

[0139] In step S40, the microfluidic chip 100 is rotated so that the liquid 200 forms a plurality of micro-droplets 201 at positions corresponding to the suction points.

[0140] It should be understood that the order of steps S20 and S30 is not limited to performing step S20 before starting step S30. In certain cases, it is also possible to perform step S30 before performing step S20.

[0141] In the above microdroplet generating method, the liquid 200 is put into the fluid passage layer 101, and the liquid 200 can be controlled to flow in the fluid passage layer 101 by using centrifugal force by rotating the microfluidic chip 100. When the liquid 200 passes through the suction point 200, the suction action of the suction point leaves a microdroplet 201 at a position corresponding to the suction point in the fluid passage layer 101. Using the above microdroplet generating method, a large amount of microdroplets 201 can be generated in a short time, the droplet generation time is greatly shortened, and the operation process is simple and convenient. Since equipment such as a high-precision micropump is not required, the system cost can be reduced. In addition, the method is scalable, and more microdroplets and multiple sets of samples can be separated by expanding the size of the microfluidic chip 100.

[0142] Specifically, the suction point can be formed in different ways. The micro-droplet generating method will be described in detail below.

[0143] In the second embodiment of the present application, the attraction point is formed by an electrode 241 of the electrode layer 23 that is turned on, and adjacent turned-on electrodes 241 are separated by electrodes 242 that are not turned on.

[0144] Correspondingly, with reference to FIG. 12, the above micro-droplet generating method includes the following steps:

[0145] Step S100 provides a microfluidic chip 100. The microfluidic chip 100 comprises an upper electrode plate 10 and a lower electrode plate 20. The upper electrode plate 10 comprises an upper cover 11, a conductive layer 12 and a first hydrophobic layer 13, which are stacked in order. The lower electrode plate comprises a second hydrophobic layer 21, a dielectric layer 22 and an electrode layer 23, which are stacked in order. The electrode layer 23 comprises a plurality of electrodes 24 arranged in an array. A fluid passage layer 101 is formed between the first hydrophobic layer 13 and the second hydrophobic layer 21.

[0146] In S200, a plurality of electrodes 24 of the electrode layer 23 are turned on to form the attraction point at the turned-on electrode 241, and the adjacent turned-on electrode 241 is separated from the other by an electrode 242 that is not turned on.

[0147] In step S300, the liquid 200 is injected into the fluid passage layer 101.

[0148] In step S400, the microfluidic chip 100 is rotated so that the liquid 200 forms a plurality of micro-droplets 201 at the turned-on electrodes 24.

[0149] It should be understood that the order of step S20 and step S30 is not limited, and step S20 may be performed before step S30, or step S30 may be performed before step S20.

[0150] In the above microdroplet generating method, the liquid 200 is put into the fluid passage layer 101, and the microfluidic chip 100 is rotated to use centrifugal force to control the liquid 200 to form a plurality of microdroplets 201 at the positions of the plurality of electrodes 24 turned on in the fluid passage layer 101. Using the above microdroplet generating method, a large amount of microdroplets 201 can be generated in a short time, the droplet generation time is greatly shortened, and the operation process is simple and convenient. Since no equipment such as a high-precision micropump is required, the system cost can be reduced. In addition, the method is scalable, and more microdroplets and multiple sets of samples can be separated by expanding the size of the microfluidic chip 100.

[0151] It should be understood that when generating the microdroplets 201, all the electrodes 24 of the electrode layer 23 are not turned on, including the turned on electrodes 241 and the turned off electrodes 242. To prevent the microdroplets 201 from bonding together, adjacent turned on electrodes 241 are separated by a turned off electrode 242. Optimally, adjacent turned on electrodes 241 are separated by two turned off electrodes 242.

[0152] It is especially important to mention that the step of injecting the liquid 200 into the fluid passage layer 101 refers to injecting the liquid 200 into the center of the fluid passage layer 101. Refer to FIG. 9(A), that is, a liquid injection hole 132 is provided in the center of the microfluidic chip 100, and the liquid 200 is injected into the fluid passage layer 101 through the liquid injection hole 132. It should be understood that the liquid 200 can also be injected into other positions of the microfluidic chip 100. The entire fluid passage layer 101 is filled, and the microfluidic chip 100 is rotated to drain the excess liquid 200. Needless to say, the liquid 200 is injected into the center of the microfluidic chip 100, and the rotation of the microfluidic chip 100 can disperse the liquid 200 from the center to the periphery, forming small liquids 200 at the turned-on electrodes 241. In this way, the amount of liquid 200 used can be effectively reduced.

[0153] It should be noted that in the above step S400, the rotation of the microfluidic chip 100 is stopped after the excess liquid 200 is discharged from the fluid passage layer 101. Specifically, referring to FIG. 9(B), liquid discharge holes 133 are provided at the four corners of the microfluidic chip 100, and the excess liquid 200 is discharged from the fluid passage layer 101 through the liquid discharge holes 133.

[0154] In this embodiment of the present application, the rotation speed of the microfluidic control 100 is greater than 0 rpm and less than or equal to 1000 rpm.

[0155] In this embodiment of the present application, the distance h between the first hydrophobic layer 13 and the second hydrophobic layer 21 is 5 μm to 600 μm.

[0156] In this embodiment of the present application, the electrode 24 is a regular hexagon, and the side length of the electrode 24 is 50 μm to 2 mm. It should be understood that the shape of the electrode 24 may be any shape or combination of shapes. The volume of the microdroplet 201 can be precisely adjusted by adjusting the size of the electrode 24, the gap distance between the electrodes 24, etc.

[0157] In this embodiment of the present application, a glass base may be selected as the material of the upper cover 11. The thickness of the upper cover 11 is 0.05 mm to 1.7 mm.

[0158] In this embodiment of the present application, an ITO conductive layer may be selected as the material of the conductive layer 12. The thickness of the conductive layer 12 is 10 nm to 500 nm.

[0159] In this embodiment of the present application, a fluorine-containing hydrophobic coating may be selected as the material of the first hydrophobic layer 13. The thickness of the first hydrophobic layer 13 is 10 nm to 200 nm.

[0160] In this embodiment of the present application, a fluorine-containing hydrophobic coating may be selected as the material of the second hydrophobic layer 21. The thickness of the second hydrophobic layer 21 is 10 nm to 200 nm.

[0161] In this embodiment of the present application, the material of the dielectric layer 22 may be selected as an organic insulating layer or an inorganic insulating layer, and the thickness of the dielectric layer 22 is 50 nm to 1000 nm.

[0162] In the second embodiment of the present application, the attraction points may be formed from hydrophilic points 131. Specifically, the upper electrode 10 has a hydrophilic point array formed on the side where the first hydrophobic layer 13 is away from the conductive layer 12, and the hydrophilic points 131 of the hydrophilic point array are the attraction points, and are spaced apart from the adjacent hydrophilic points 131.

[0163] Correspondingly, as shown in FIG. 13, the above micro-droplet generating method includes the following steps:

[0164] Step S1000 provides a microfluidic chip 100. The microfluidic chip 100 comprises an upper electrode plate 10 and a lower electrode plate 20. The upper electrode plate 10 comprises an upper cover 11, a conductive layer 12 and a first hydrophobic layer 13, which are stacked in order. The lower electrode plate comprises a second hydrophobic layer 21, a dielectric layer 22 and an electrode layer 23, which are stacked in order. The electrode layer 23 comprises a plurality of electrodes 24 arranged in an array. A fluid passage layer 101 is formed between the first hydrophobic layer 13 and the second hydrophobic layer 21.

[0165] In step S2000, a hydrophilic point 131 is formed in the first hydrophobic layer 13, and the hydrophilic point 131 is disposed at the suction point and separated from the adjacent hydrophilic points 131.

[0166] In step S3000, the liquid 200 is injected into the fluid passage layer 101.

[0167] In step S4000, the microfluidic chip 100 is rotated so that the liquid 200 forms a plurality of micro-droplets 201 at the turned-on electrodes 24.

[0168] In the above microdroplet generating method, the liquid 200 is put into the fluid passage layer 101, and the liquid 200 can be controlled to flow in the fluid passage layer 101 by using centrifugal force by rotating the microfluidic chip 100. When a large droplet passes through the hydrophilic point 131, the hydrophilic effect of the hydrophilic point 131 leaves the microdroplet 201 at the position corresponding to the hydrophilic point 131 in the fluid passage layer 101. Using the above microdroplet generating method, a large amount of microdroplets 201 can be generated in a short time, the droplet generation time is greatly shortened, and the operation process is simple and convenient. In the above microdroplet generating method, the microdroplets 201 can be separated without controlling the electrodes 24, making the operation easier. Since equipment such as a high-precision micropump is not required, the system cost can be reduced. In addition, the method is scalable, and more microdroplets and multiple sets of samples can be separated by expanding the size of the microfluidic chip 100.

[0169] It should be understood that the step of injecting the liquid 200 into the fluid passage layer 101 refers to injecting the liquid 200 into the center of the fluid passage layer 101. That is, a liquid injection hole 132 is provided in the center of the microfluidic chip 100, and the liquid 200 is injected into the fluid passage layer 101 through the liquid injection hole 132. It should be understood that the liquid 200 can also be injected into other positions of the microfluidic chip 100. The entire fluid passage layer 101 is filled, and the microfluidic chip 100 is rotated to drain the excess liquid 200. Needless to say, the liquid 200 is injected into the center of the microfluidic chip 100, and the rotation of the microfluidic chip 100 can disperse the liquid 200 from the center to the periphery, forming small liquids 200 at the turned-on electrodes 241. In this way, the amount of liquid 200 used can be effectively reduced.

[0170] In this embodiment of the present application, in step S4000, the rotation of the microfluidic chip 100 is stopped after the excess liquid 200 is discharged from the fluid passage layer 101. Specifically, liquid discharge holes 133 are provided at the four corners of the microfluidic chip 100, and the excess liquid 200 is discharged from the fluid passage layer 101 through the liquid discharge holes 133.

[0171] In this embodiment of the present application, the rotation speed of the microfluidic control 100 is greater than 0 rpm and less than or equal to 1000 rpm.

[0172] In this embodiment of the present application, the distance between the first hydrophobic layer 13 and the second hydrophobic layer 21 is 5 μm to 600 μm. That is, the gap h of the fluid passage layer 101 is 5 μm to 600 μm.

[0173] In this embodiment of the present application, the method of generating the hydrophilic dots 131 is as follows: using laser or plasma to process the hydrophobic coating at the target position of the first hydrophobic layer 13, the hydrophilic dots 131 are formed.

[0174] In this embodiment of the present application, the multiple hydrophilic points 131 of the first hydrophobic layer 13 are arranged in an array.

[0175] It should be understood that in the second embodiment, the micro-droplet generating system uses the centrifugal force obtained by the rotation of the rotary drive unit to realize an operation similar to spin coating on the surface of the electrode array, and generates normal liter-level droplets at high throughput through hydrophilic modification of the electrodes 24 or the upper cover 11 in an array format. The volume of the droplets can be precisely adjusted by adjusting the size of the electrodes 24, the gap distance, or the size of the hydrophilic modification points.

[0176] Third embodiment 14 to 21, the detailed structure of a micro-droplet generating system and a micro-droplet generating method according to the third embodiment of the present application are specifically described. The third embodiment is a modified example of the first embodiment.

[0177] The micro-droplet generating system of the third embodiment includes a micro-fluidic chip 100 and a droplet driving unit connected to the micro-fluidic chip 100. The micro-fluidic chip 100 includes an upper electrode plate 10 and a lower electrode plate 20, and the upper electrode plate 10 includes an upper cover 11, a conductive layer 12 and a first hydrophobic layer 13, which are arranged in sequence. The lower electrode plate 20 includes a second hydrophobic layer 21, a dielectric layer 22 and an electrode layer 23, which are arranged in sequence. The first hydrophobic layer 13 and the second hydrophobic layer 21 are arranged opposite to each other, and the fluid passage layer 101 is formed between the first hydrophobic layer 13 and the second hydrophobic layer 21, and the electrode layer 23 includes a plurality of electrodes 24 arranged in an array. At least one of the upper electrode plate 10 and the lower electrode plate 20 forms a plurality of suction points, which are used to suck in the liquid 200. The droplet driving unit is used to drive the liquid 200 poured into the fluid passage layer 101 to flow in the fluid passage layer 101, thereby forming a micro-droplet 201 at the position of the suction point.

[0178] Specifically, as shown in Figures 14 and 15, the difference from the first embodiment is that the microfluidic chip 100 is provided with a first sample inlet hole 134 and a first sample outlet hole 135, and the first sample inlet hole 134 and the first sample outlet hole 135 are provided on a first diagonal line of the microfluidic chip 100. The droplet driving unit includes a first micropump 41 and a third micropump 43, and the first micropump 41 is connected to the first sample inlet hole 134 and is used to inject liquid 200 into the fluid passage layer 101. In this way, the fluid passage layer 101 is filled with the liquid 200. The third micropump 43 is connected to the first sample outlet hole 135 and is used to extract the liquid 200 discharged from the first sample outlet hole 135.

[0179] It should be noted that the first sample inlet hole 134 and the first sample outlet hole 135 are arranged diagonally in order to allow the liquid 200 to fill the fluid passage layer 101 without leaving any air bubbles.

[0180] Further, the microfluidic chip 100 is provided with a second sample inlet hole 136 and a second sample outlet hole 137, the second sample inlet hole 136 and the second sample outlet hole 137 are provided on a second diagonal of the microfluidic chip 100, and the droplet driving unit includes a second micropump 42 and a fourth micropump 44, the second micropump 42 is connected to the second sample inlet hole 136 and is used to inject a medium 300 into the fluid passage layer 101. When the second micropump 42 injects a medium into the fluid passage layer 101, the liquid 200 at a position other than the suction point is pushed into the medium 300, and the liquid 200 leaves a microdroplet 201 at a position corresponding to the suction point, and the medium 300 encapsulates the microdroplet. The fourth micropump 44 is connected to the second sample outlet hole 137 and is used to extract the medium 300 coming out of the second sample outlet hole 137.

[0181] It is important to note that the first sample inlet hole 134 and the first sample outlet hole 135 are diagonally arranged to ensure that the medium 300 can fully drain the liquid 200 that is not the suction point of the entire fluid passage layer 101. The medium 300 can be air, oil, or other medium 300.

[0182] It should be noted that the first micro pump 41, the second micro pump 42, the third micro pump 43 and the fourth micro pump 44 are digital injection pumps, but are not limited to digital pumps. They can be used as long as they can guarantee that the liquid 200 flows in and out stably.

[0183] In this embodiment of the present application, the material of the upper cover can be glass-based, and the thickness of the upper cover can be 0.05mm-1.7mm.

[0184] In this embodiment of the present application, the material of the conductive layer 12 can be an ITO conductive layer, and the thickness of the conductive layer 12 can be 10 nm-1000 nm.

[0185] In this embodiment of the present application, the thickness of the first hydrophobic layer 13 may be 10 nm-200 nm.

[0186] In this embodiment of the present application, the thickness of the second hydrophobic layer 21 may be 10 nm-200 nm.

[0187] In this embodiment of the present application, the material of the dielectric layer 22 can be an organic or inorganic insulating material, and the thickness of the dielectric layer 22 can be 50 nm-1000 nm.

[0188] In this embodiment of the present application, the material of the electrode layer 23 can be a metal or its oxide conductive material, and the thickness of the electrode layer 23 can be 10 nm-500 nm.

[0189] In this embodiment of the present application, the lower plate may include a substrate 25. The substrate 25 is disposed on the side of the electrode layer 23 that is far from the dielectric layer 22. The substrate 25 is used to protect the lower plate 20. In one embodiment, the material of the substrate 25 may be glass or PCB plate material. The thickness of the substrate 25 may be 0.05mm-5mm.

[0190] It should be understood that an attraction point can be formed on the upper plate 10, an attraction point can be formed on the lower plate 20, or an attraction point can be simultaneously formed on the upper plate 10 and the lower plate 20. A plurality of attraction points on the upper plate 10 or the lower plate 20 are arranged in an array.

[0191] The attraction points can also be formed by hydrophilic points 131. Specifically, the upper electrode 10 has a hydrophilic point array formed on the side of the first hydrophobic layer 13 far from the conductive layer 12, and the hydrophilic points 131 of the hydrophilic point array are the attraction points, and adjacent hydrophilic points 131 are spaced apart. More specifically, the first hydrophobic layer 13 is hydrophilically modified, for example, by using micro-nano processing techniques such as photolithography and etching, and the first hydrophobic layer 13 is treated with hydrophobic coating at required positions, thereby forming a hydrophilic point array.

[0192] 16 shows the liquid injection process of the above-mentioned micro-droplet generating system. By adjusting the first micro-pump 41, the liquid 200 flows in through the first sample injection hole 134, while the third micro-pump 43 extracts the excess air. When the micro-fluidic chip 100 is filled with the injected liquid 200, the excess liquid is discharged through the first sample discharge hole 135. The whole process ensures the pressure equalization inside the micro-fluidic chip 100, and the entire fluid passage layer is filled with the liquid 200, completing the liquid injection.

[0193] FIG. 17 shows the sample ejection process of the micro-droplet generating system, that is, the process of forming high-density micro-droplets. First, the electrode 24 of the micro-fluidic chip 100 that needs to generate the micro-droplets 201 is powered. Usually, the generated high-density micro-droplets 201 are separated from each other by one electrode 24 to prevent crosstalk between each other. That is, the powered electrode 24 is separated from the non-powered electrode 24. By adjusting the second micro-pump 42, the medium 300 is injected into the micro-fluidic chip 100 from the second sample injection hole 136. At the same time, the fourth micro-pump 44 is used to extract the liquid 200. After the liquid medium 200 is completely ejected from the second sample ejection hole 137, the remaining medium 300 is ejected from the second sample injection hole, thus completing the sample ejection. The micro-droplet 201 is left at the position of the powered electrode 24 in the micro-fluidic chip 100. At the same time, the micro-droplet 201 is enveloped in the target medium.

[0194] Figures 18 and 19 show the process of the digital Elisa operation of the microdroplet generation system. As shown in Figure 18, the mixed solution contains microspheres (magnetic beads, PS, etc.), capture antibodies 52, target antigens and fluorescently labeled antibodies 54. After the mixed solution 50 undergoes an immune reaction, a first microsphere 511 containing the target antigen and fluorescently labeled antibodies and a second microsphere 512 not containing the target antigen and fluorescently labeled antibodies are generated. Next, the microspheres 51 are washed to remove non-specifically bound proteins. Next, a substrate is added, and finally, the mixed solution is pumped into the electrowetting microarray microfluidic chip 100 using the above-mentioned microdroplet generation method to form a high-density microdroplet array in which each droplet contains only one or more microspheres 51. Figure 19 shows a cross-sectional view of the electrowetting microfluidic chip 100 in which microdroplets 201 are generated. Among them, the microsphere 51 containing the target antigen 53 is fluorescently labeled with a fluorescent antibody 54, which emits fluorescence and is digitized by a CCD imaging system, and the target protein concentration is calculated based on the Poisson distribution theory. This calculation method is not a traditional Elisa simulation calculation, but a digitalized calculation, so it is called digital Elisa.

[0195] Additionally, by marking different fluorescent antibodies 54 with fluorescent labels of different absorption and emission wavelengths, detection of multiple target antigens 53 is possible.

[0196] This method uses the standard enzyme-linked immunosorbent assay (Elisa) to enable quantitative detection of extremely small amounts of protein. The notable feature of this method is that it realizes single molecule detection and uses simulation calculations, making the detection sensitivity much higher than traditional methods. Although the detection principle is similar to that of Quanterix, the high-density array micro-droplet formation method is completely different. What is different from Quanterix is ​​that the above micro-droplet generation method uses a technology called electrowetting to form a high-density droplet array, allowing the generated droplets to be controlled at will.

[0197] In the above micro-droplet generating system, the first micro-pump 41 pours the liquid 200 into the fluid passage layer 101, so that the liquid 200 fills the fluid passage layer 101. The liquid 200 is sucked into the powered electrode 24. The second micro-pump 42 pours the medium 300 into the fluid passage layer 101, so that the liquid 200 at the non-suction point is pushed into the medium 300, so that the liquid 200 forms a plurality of micro-droplets 201 at the powered electrode 24 in the fluid passage layer 101, and the medium 300 envelops the micro-droplets 201. Using the above micro-droplet generating method, a large amount of micro-droplets 201 can be generated quickly, the droplet generation time can be greatly shortened, and the operation process is simple and convenient.

[0198] It should be understood that by adjusting the gap of the fluid passage layer 101 and the size of the electrodes 24, the volume of the micro-droplets 201 can be precisely controlled from fL to μL. By adjusting the density of the electrodes 24 and the size of the micro-fluidic chip 100, the number of micro-droplets 201 can be controlled. After the separation of the high-density nL droplets is completed, the droplets can be precisely controlled by the digital micro-fluidic chip to perform experiments and testing, such as ddPCR, dLAMP, and dELISA single-cell experiments.

[0199] In this system, after the high-density micro-droplets complete the corresponding experiment, the micro-fluidic chip 100 can be quickly washed by injecting a washing solution into the fluid passage layer 101 with a micro-pump. The micro-fluidic chip 100 can be recycled. By controlling the digital micro-pump, the medium 300 or the washing solution is injected into the sample injection hole, and at the same time, the waste liquid in the micro-fluidic chip 100 is discharged from the sample discharge hole. This method is fast, convenient and easy to operate.

[0200] As shown in Figure 20, a method for generating micro droplets is presented in the third embodiment, which includes the following steps:

[0201] Step S61: Provide a microfluidic chip 100. The microfluidic chip 100 comprises an upper electrode plate 10 and a lower electrode plate 20. A fluid passage layer 101 is formed between the upper electrode plate 10 and the lower electrode plate 20.

[0202] In step S62, at least one of the upper plate 10 and the lower plate 20 forms a plurality of suction points, which are used to suck the liquid 200.

[0203] In step S63, the liquid 200 is poured into the fluid passage layer 101 so that the fluid passage layer 101 is filled with the liquid 200.

[0204] In step S64, the medium 300 is poured into the fluid passage layer 101, and the liquid 200 at the non-suction points is pushed into the medium 300, so that the liquid 200 leaves the micro-droplets 201 at the positions corresponding to the suction points, and the medium 300 envelops the micro-droplets 201.

[0205] It should be understood that the order of steps S62 and S63 is not limited to performing step S62 before starting step S63. In a specific case, it is also possible to perform step S62 after performing step S63.

[0206] As shown in FIG. 21, the above microdroplet generation method includes the following steps:

[0207] Step S610 provides a microfluidic chip 100. The microfluidic chip 100 comprises an upper electrode plate 10 and a lower electrode plate 20. The upper electrode plate 10 comprises an upper cover 11, a conductive layer 12 and a first hydrophobic layer 13, which are stacked in order. The lower electrode plate comprises a second hydrophobic layer 21, a dielectric layer 22 and an electrode layer 23, which are stacked in order. The electrode layer 23 comprises a plurality of electrodes 24 arranged in an array. A fluid passage layer 101 is formed between the first hydrophobic layer 13 and the second hydrophobic layer 21.

[0208] In step S620, the liquid 200 is poured into the fluid passage layer 101 so that the liquid 200 fills the fluid passage layer 101.

[0209] In step S630, a plurality of electrodes 24 in the electrode layer 23 are turned on, so that adjacent turned-on electrodes 241 are spaced apart by non-turned-on electrodes 242, and the turned-on electrodes 241 form attraction points.

[0210] In step S640, the medium 300 is poured into the fluid passage layer 101, and the liquid 200 at the non-suction points is pushed into the medium 300, so that the liquid 200 leaves the micro-droplets 201 at the positions corresponding to the suction points, and the medium 300 envelops the micro-droplets 201.

[0211] It should be understood that the order of steps S620 and S630 is not limited, and step S620 may be performed before step S630, or step S630 may be performed before step S620.

[0212] It should be understood that when generating the microdroplets 201, all the electrodes 24 of the electrode layer 23 are not turned on, including the turned on electrodes 241 and the turned off electrodes 242. To prevent the microdroplets 201 from bonding together, adjacent turned on electrodes 241 are separated by a turned off electrode 242. Optimally, adjacent turned on electrodes 241 are separated by two turned off electrodes 242.

[0213] It should be understood that in the third embodiment, the present application uses a digital syringe pump to inject a sample into the digital microfluidic chip at a certain volume and flow rate, achieving a control similar to dispensing. The digital syringe pump then ejects the sample, and leaves a high-density droplet array at the powered position of the electrodes 24 by controlling the electrodes 24. The volume of the droplets can be precisely adjusted by adjusting the number of electrodes 24, the size of the electrodes 24, the gap distance, etc.

[0214] Fourth embodiment As shown in Figures 22 to 27, the detailed structure of the micro-droplet generating system and the micro-droplet generating method of the fourth embodiment of the present application are specifically described. As shown in Figures 22 to 24, in the fourth embodiment, the micro-droplet generating system includes a microfluidic chip 100 consisting of an upper electrode 10 and a lower electrode 20. The fluid passage layer 101 is formed between the upper electrode 10 and the lower electrode 20, and at least one of the upper electrode 10 and the lower electrode 20 forms a plurality of suction points, which are used to suck the liquid 200. An angle is set between the plane on which the upper electrode 10 is located and the plane on which the lower electrode 20 is located. The upper electrode 10 is provided with a plurality of sample injection holes, which are located at the ends of the upper electrode 10, and which are used to inject the liquid 200. The fluid passage layer 101 includes a first end and a second end that are arranged opposite to each other. The height of the first end of the fluid passage layer 101 is lower than the height of the second end of the fluid passage layer 101, and when liquid 200 is injected into the first end of the fluid passage layer 101 through the sample injection hole, the liquid 200 is subjected to surface tension and moves from the first end to the second end, forming a microdroplet 201 at the suction point.

[0215] It should be understood that the height of the first end of the fluid passage layer 101 is lower than the height of the second end of the fluid passage layer 101 because the distance between the upper electrode plate 10 and the lower electrode plate 20 at the first end is the smallest, and the distance between the upper electrode plate 10 and the lower electrode plate 20 at the second end is the largest.

[0216] In particular, the angle between the upper electrode plate 10 and the lower electrode plate 20 is greater than 0° and less than 3°. At the first end, the distance between the upper electrode plate 10 and the lower electrode plate 20 is 0 μm to 200 μm.

[0217] As shown in Figures 22 to 24, the upper plate 10 includes an upper cover 11, a conductive layer 12 and a first hydrophobic layer 13, which are arranged in sequence. The lower plate 20 includes a second hydrophobic layer 21, a dielectric layer 22 and an electrode layer 23, which are arranged in sequence. The first hydrophobic layer 13 and the second hydrophobic layer 21 are arranged opposite to each other, and a fluid passage layer 101 is formed between the first hydrophobic layer 13 and the second hydrophobic layer 21. The electrode layer 23 includes a plurality of electrodes 24 arranged in an array.

[0218] As shown in Figures 22 to 24, in this application, a spacer is used to raise one side of the upper electrode plate 10, to create a certain angle between the upper electrode plate 10 and the lower electrode plate 20, and the distance between the upper electrode plate 10 and the lower electrode plate 20 is changed. The distance between the upper electrode plate 10 and the lower electrode plate 20 gradually increases from right to left. Referring to Figures 23 and 24, when a droplet is injected into the microfluidic chip 100 from the right side, the liquid 200 moves to the area with a larger gap. That is, it moves from right to left. Here, a voltage is applied to the electrode layer 23, and the surface of the corresponding electrode 24 becomes hydrophilic. When the liquid 200 flows through the electrode 24 to which the voltage is applied, it splits into multiple microdroplets 201 with the size of a single electrode 24 and is discharged from the inside. The microdroplets 201 are separated from each other by several turned-on electrodes 241 to prevent the microdroplets 201 from fusing with each other. The faster the liquid 200 is injected, the more successful it will be in breaking up into micro-droplets 201 .

[0219] FIG. 25 is a bird's-eye view of the droplet movement, showing the process of the micro-droplet generation method of the micro-droplet generation system. In this embodiment of the present application, the angle formed by the upper cover 11 and the surface of the electrode 24 is used to force the large droplets to move to the larger gap, and then the direction of the large droplets is controlled by electrowetting to generate normal liter-level droplets over the entire surface of the attraction point area. The volume of the droplets can be controlled by adjusting the size of the electrode 24, the gap distance, and the size of the hydrophilic modification point. That is, the micro-droplet generation system can generate a large number of micro-droplets at a high speed. In addition, a large number of micro-droplets 201 with different volumes can be generated through calculation, which is convenient for generating samples with different concentrations.

[0220] In traditional digital microfluidic control, a single microdroplet 201 is generated from a larger droplet, and the microdroplet 201 is controlled to be transported to a corresponding position. In the above microdroplet generation method, liquid 200 is injected into a first end of the fluid passage layer 101, and the injected liquid 200 is subjected to surface tension, so that the liquid 200 gradually moves from the first end to the second end, that is, in the direction of the arrows shown in Figures 22 to 24. The microdroplet 201 is left in the fluid passage layer 101 at a position corresponding to the suction point, and the time for droplet generation can be greatly shortened.

[0221] In future experiments, the desired droplet volume can be selected for the experiment. After the high-throughput normal liter droplets are separated, the microfluidic chip 100 can be used for related experiments and testing, such as ddPCR, dLAMP, and dELISA single-cell experiments. It can also be used for other nucleic acid tests, such as isothermal amplification. In addition, the microfluidic chip 100 can be used to narrow down or perform independent experiments on any of the microdroplets, and the size of the microfluidic chip 100 can be expanded to separate more microdroplets or more groups of samples.

[0222] It is particularly important to note that the shape of the electrodes 24 can be either hexagonal or square. Of course, the shape of the electrodes 24 is not limited to hexagonal or square. Specifically, the electrode layer 23 is an n×m electrode array, where n and m are both positive integers.

[0223] In this embodiment of the present application, the shape of the electrode 24 is a square. The side length ranges from 50 μm to 2000 μm. It should be understood that the shape of the electrode 24 can be any shape and any combination of shapes.

[0224] It should be understood that the volume of the microdroplet 201 can be precisely adjusted by adjusting the size of the electrodes 24, the gap distance between multiple electrodes 24, etc. By adjusting different electrode 24 sizes, single droplets of different volumes can be rapidly generated.

[0225] In this embodiment of the present application, the material of the upper cover 11 may be glass-based, and the thickness of the upper cover 11 is 0.7mm-1.7mm.

[0226] In this embodiment of the present application, an ITO conductive layer may be selected as the material of the conductive layer 12. The thickness of the conductive layer 12 may be 10 nm to 500 nm.

[0227] In this embodiment of the present application, a fluorine-containing hydrophobic coating may be selected as the material of the first hydrophobic layer 13. The thickness of the first hydrophobic layer 13 may be 10 nm to 200 nm.

[0228] In this embodiment of the present application, a fluorine-containing hydrophobic coating may be selected as the material of the second hydrophobic layer 21. The thickness of the second hydrophobic layer 21 may be 10 nm to 200 nm.

[0229] In this embodiment of the present application, the material of the dielectric layer 22 may be selected as an organic insulating layer or an inorganic insulating layer, and the thickness of the dielectric layer 22 may be 50 nm to 1000 nm.

[0230] In this embodiment of the present application, the material of the electrode layer 23 may be selected as transparent conductive glass or metal, and the thickness of the electrode layer 23 may be 10 nm-1000 nm.

[0231] It should be understood that it is possible to form an attraction point on the upper plate 10 and also to form an attraction point on the lower plate 20, or to form an attraction point on both the upper plate 10 and the lower plate 20 at the same time.

[0232] Specifically, the attraction points can be formed using different methods.

[0233] In this embodiment of the present application, the attraction point can be formed at an electrode 241 in the electrode layer 23 that is turned on, and adjacent turned-on electrodes 241 are spaced apart by electrodes 242 that are not turned on.

[0234] The attraction points can also be formed by hydrophilic points 131. Specifically, the upper electrode 10 has a hydrophilic point array formed on the side of the first hydrophobic layer 13 far from the conductive layer 12, and the hydrophilic points 131 of the hydrophilic point array are the attraction points, and the adjacent hydrophilic points 131 are spaced apart. More specifically, the first hydrophobic layer 13 employs laser or plasma to process hydrophobic coating at required positions, thereby forming a hydrophilic point array.

[0235] As shown in FIG. 26, the fourth embodiment of the micro-droplet generating system includes the following steps.

[0236] Step S51: providing a microfluidic chip 100. The microfluidic chip 100 comprises an upper electrode plate 10 and a lower electrode plate 20. An angle is set between a plane on which the upper electrode plate 10 is located and a plane on which the lower electrode plate 20 is located. A fluid passage layer 101 is formed between the upper electrode plate 10 and the lower electrode plate 20. A plurality of sample injection holes are provided in the upper electrode plate, and the sample injection holes are located at the ends of the upper electrode plate 10, and the sample injection holes are used to inject samples. The fluid passage layer 101 includes a first end and a second end that are disposed opposite to each other. The height of the first end of the fluid passage layer 101 is lower than the height of the second end of the fluid passage layer 101.

[0237] In step S52, at least one of the upper plate 10 and the lower plate 20 forms a plurality of suction points, which are used to suck the liquid 200.

[0238] Step S53: Inject liquid 200 into the first end of the fluid passage layer 101 through the sample injection hole.

[0239] Step S54: When the liquid 200 is injected into the fluid passage layer 101, the liquid 200 moves from the first end to the second end due to surface tension, and the liquid 200 forms a micro-droplet 201 at a position corresponding to the suction point.

[0240] Specifically, step 54 involves injecting the liquid 200 into the fluid passage layer 101, gradually drawing the upper plate 10 and the lower plate 20 closer to each other, and the liquid 200 moving from the first end to the second end under the effect of surface tension, forming micro-droplets 201 at positions corresponding to the attraction points.

[0241] It should be understood that the order of steps S52 and S53 is not limited to performing step S52 before starting step S53. In a specific case, step S53 may be performed before performing step S54.

[0242] As shown in FIG. 26, the fourth embodiment of the micro-droplet generating system includes the following steps.

[0243] Step S510 provides a microfluidic chip 100. The microfluidic chip 100 includes an upper electrode plate 10 and a lower electrode plate 20. An angle is set between the plane on which the upper electrode plate 10 is located and the plane on which the lower electrode plate 20 is located. The upper electrode plate 10 includes an upper cover 11, a conductive layer 12 and a first hydrophobic layer 13 arranged in sequence. The lower electrode plate 20 includes a second hydrophobic layer 21, a dielectric layer 22 and an electrode layer 23 arranged in sequence. The electrode layer 23 includes a plurality of electrodes 24 arranged in an array. A fluid passage layer 101 is formed between the first hydrophobic layer 13 and the second hydrophobic layer 21. The fluid passage layer 101 includes a first end and a second end arranged opposite to each other. The height of the first end of the fluid passage layer 101 is lower than the height of the second end of the fluid passage layer 101. A number of sample injection holes are provided in the upper plate 10, which are located at the edge of the upper plate 10 and are used for sample injection.

[0244] In step S520, the liquid 200 is injected into the first end of the fluid passage layer 101.

[0245] In this embodiment of the present application, liquid 200 is injected into the first end of fluid passage layer 101 at a sample injection hole.

[0246] In step S530, a plurality of electrodes 24 in the electrode layer 23 are turned on, and adjacent turned-on electrodes 241 are spaced apart by non-turned-on electrodes 242.

[0247] In step S540, the upper electrode plate 10 and the lower electrode plate 20 are gradually brought closer to each other, and the liquid 200 moves from the first end to the second end. The liquid 200 forms micro-droplets 201 at positions corresponding to the suction points.

[0248] It should be understood that the order of steps S520 and S530 is not limited, and step S520 may be performed first before step S530, or step S530 may be performed first before step S520.

[0249] In the above-mentioned micro-droplet generating method, liquid 200 is injected into a first end of the fluid passage layer 101, and when the upper electrode plate 10 and the lower electrode plate 20 are brought closer to each other, the liquid 200 moves from the first end to the second end. When the liquid 200 passes through the turned-on electrodes 24, the liquid 200 forms a plurality of micro-droplets 201 in the fluid passage layer 101 at the positions of the turned-on electrodes 24.

[0250] Using the above-mentioned micro-droplet generating method, a large amount of micro-droplets 201 can be generated in a short time, the droplet generating time can be greatly shortened, and the operation process is simple and convenient. Since equipment such as a high-precision micropump is not required, the system cost can be reduced. In addition, the method is scalable, and more micro-droplets and multiple sets of samples can be separated by expanding the size of the microfluidic chip 100.

[0251] It should be understood that when generating the microdroplets 201, not all of the electrodes 24 of the electrode layer 23 are turned on, including the turned on electrodes 241 and the turned off electrodes 242. To prevent the microdroplets 201 from bonding together, adjacent turned on electrodes 241 are separated by an unturned electrode 242. It should be understood that adjacent electrodes 241 are separated by at least one unturned electrode 242. Optimally, adjacent turned on electrodes 241 are separated by two unturned electrodes 242. It is particularly noted that in the step of injecting the liquid 200 into the first end of the fluid passage layer 101, the injection speed of the liquid 200 is 1 μL / s-10 μL / s.

[0252] In the above microdroplet generating method, the liquid 200 is injected into the first end of the fluid passage layer 101, and when the upper electrode 10 and the lower electrode 20 approach each other, the liquid 200 moves from the first end to the second end. When the liquid 200 passes through the suction point, the suction action of the suction point leaves the microdroplet 201 at the position corresponding to the suction point in the fluid passage layer 101. Using the above microdroplet generating method, a large amount of microdroplets 201 can be generated in a short time, the droplet generation time is greatly shortened, and the operation process is simple and convenient. Since no equipment such as a high-precision micropump is required, the system cost can be reduced. In addition, the method is scalable, and more microdroplets and multiple sets of samples can be separated by expanding the size of the microfluidic chip 100.

[0253] In the above microdroplet generation method, multiple microdroplets 201 can be generated simultaneously and quickly by combining electrowetting with changing the gap distance between the upper electrode plate 10 and the lower electrode plate 20. In addition, the volume of the microdroplets 201 can be controlled by adjusting the gap between the upper electrode plate 10 and the lower electrode plate 20 and the size of the electrode 24. The operation method is convenient and easy to control. At the same time, the droplets can be controlled to move spontaneously and leave the microdroplets 201 in a specified place or area. Furthermore, by controlling the on and off of the electrode 24, the microdroplets 201 can be moved, and the droplets can be controlled through electrowetting to complete the experiment on the chip. It can be applied to multiple biochemical applications of droplets.

[0254] Actual tests have shown that the above-mentioned micro-droplet generating method can separate a large number of droplets at high speed and control the movement of the separated droplets, thereby improving the separation efficiency.

[0255] Fifth embodiment As shown in Figures 28 to 35, the detailed structure of a micro-droplet generating system and a micro-droplet generating method according to the fifth embodiment of the present application are specifically described.

[0256] Referring to FIG. 28, the micro-droplet generating system of the fourth embodiment includes the following items:

[0257] The microfluidic chip is composed of an upper electrode plate 10 and a lower electrode plate 20. The fluid passage layer 101 is formed between the upper electrode plate 10 and the lower electrode plate 20.

[0258] A number of suction points are formed on the lower electrode 20, which are used to suction liquid. The liquid sample flows through the fluid passage layer 101 and forms micro-droplets 201 at the suction points.

[0259] The bottom plate includes an electrode layer 23. The electrode layer 23 includes a plurality of electrodes 24 arranged in an array having at least two different shapes.

[0260] The attraction points are formed by the turned-on electrodes 241 of the electrode layer 23, and adjacent turned-on electrodes 241 are separated by turned-off electrodes 242.

[0261] It should be noted that in the micro-droplet generating system introduced in the embodiment of the present application, the liquid sample is put into the fluid passage layer 101, so that the liquid sample fills the fluid passage layer 101, and the liquid sample flows in the fluid passage layer, and the droplet sample forms a micro-droplet at a position corresponding to the attraction point. Specifically, by controlling the on and off of the electrodes 24 of the electrode layer 23, the electrowetting principle (when liquid exists in the electrode, the wettability of the solid-electrolyte interface at the position corresponding to the electrode can be changed by applying an electric field to the electrode. This also changes the contact angle between the droplet and the electrode interface. Due to the electric field difference between the electrodes in the droplet area, when the contact angles are different, a lateral propulsion force is generated, and the droplet moves laterally on the electrode substrate) is utilized, and the liquid sample is attracted by the turned-on electrode, and a plurality of micro-droplets are formed at positions corresponding to the plurality of turned-on electrodes in the fluid passage layer. Using this micro-droplet generating method, the droplet generation time can be greatly shortened and the droplet generation stability can be improved. In addition, the size of the generated droplets can be adjusted at any time according to needs, and the operation process is simple and convenient. Since no equipment such as a high-precision micropump is required, the system cost can be reduced. In addition, the scalability is good, and more microdroplets and multiple sets of samples can be separated through the expansion of the size of the microfluidic control. Furthermore, the electrode layer 23 of the present application includes a plurality of electrodes 24 arranged in an array of at least two different shapes. For example, the electrode layer 23 includes a plurality of electrodes 24 arranged in an array of at least two different shapes, such as square, rectangular, hexagonal, pentagonal, triangular, and circular. In this way, by controlling the on and off of the electrodes 24, it is possible to realize that a large droplet forms a microdroplet 201 with the plurality of electrodes 24 arranged in one of the arrays. In addition, a related experiment of the microdroplet is performed with the plurality of electrodes 24 arranged in an array of another shape. For example, a microdroplet is formed from a large droplet with the plurality of electrodes 24 arranged in a square array, while a related experiment of the microdroplet is performed with the plurality of electrodes 24 arranged in a circular array. In this way, the transfer of liquid samples can be avoided.

[0262] Specifically, in the above embodiment, adjacent turned-on electrodes 241 are spaced apart by an unturned electrode 242. Optimally, adjacent turned-on electrodes 241 are spaced apart by two unturned electrodes 242.

[0263] In some embodiments, the electrode layer 23 includes a plurality of square electrodes 243 arranged in an array and fluorine arranged in an array. The volume of the microdroplets can be precisely adjusted by adjusting the size of the electrodes and the gap distance between the electrodes. Different electrode sizes can be controlled to rapidly form single droplets of different volumes. For example, the volume of the microdroplets can reach the pL range by adjusting the size of the electrodes and the gap distance between the electrodes. Furthermore, the position and number of the microdroplets formed can be controlled by controlling the position and number of the electrodes that are turned on. That is, the density at which the microdroplets are formed can be precisely controlled.

[0264] Specifically, the square electrodes 243 and the hexagonal electrodes 244 can be arranged to cross each other, or can be arranged in other ways according to needs.

[0265] In the above embodiment, a plurality of hexagonal electrodes 244 arranged in an array are located between two plurality of square electrodes 243 arranged in an array. Referring to steps S1 to S4 in FIG. 30, in application, liquid 200 is present in the area corresponding to the hexagonal electrodes 244, and the liquid 200 forms microdroplets 201 by controlling the on and off of the electrodes in the hexagonal electrodes 244. Next, the microdroplets 201 move to the corresponding areas of the square electrodes 243 by controlling the on and off of the electrodes, thereby completing the droplet sorting. In addition, by performing the relevant experiment of the microdroplets in the square electrode 243 area, the transfer between the microdroplets and the large droplets can be avoided.

[0266] In some embodiments, as shown in FIG. 31 , the electrode layer 23 includes a plurality of square electrodes 243 arranged in an array and a plurality of hexagonal electrodes 244 arranged in an array located on either side of the plurality of square electrodes 243 arranged in an array. In the above embodiment, a plurality of square electrodes arranged in an array are located between two plurality of hexagonal electrodes 244 arranged in an array. With reference to steps S1 to S3 in FIG. 32, in application, the liquid 200 in the area corresponding to the hexagonal electrode 244 and the electrodes in the hexagonal electrode 244 are controlled to be on and off, so that the liquid 200 forms a microdroplet 201. Next, the electrodes are controlled to be on and off, so that the microdroplet 201 moves to the area corresponding to the square electrode 243, thereby completing the droplet sorting. In addition, by performing a related experiment of the microdroplet in the square electrode 243 area, the transfer between the microdroplet and the large droplet is avoided.

[0267] Specifically, in some embodiments, the side length of the hexagonal electrode 244 is 50 μm to 2 mm, and the side length of the square electrode 243 is 50 μm to 2 mm. In practice, the side lengths of the hexagonal electrode 244 and the square electrode 243 can be adjusted according to the needs of use.

[0268] In some embodiments, referring to FIG. 33, the electrode layer 23 includes a plurality of first square electrodes 2431 arranged in an array, a plurality of first hexagonal electrodes 2441 arranged in an array, a plurality of second hexagonal electrodes 2442 arranged in an array, and a plurality of second square electrodes 2432 arranged in an array, which are connected in series.

[0269] In the above embodiment, the electrode layer 23 includes two square electrodes arranged in an array and two hexagonal electrodes arranged in an array. The square electrodes are between the hexagonal electrodes, and the square electrodes and the hexagonal electrodes have different side lengths and sizes. Steps S1 to S9 in FIG. 33 show one specific application. A liquid 200 containing a plurality of cells 202 enters a corresponding location of the first square electrode 2431, and by controlling the on and off of the electrode, the liquid 200 containing a plurality of cells 202 moves to a corresponding location of the first hexagonal electrode 2441 to form a microdroplet 201 containing one cell 202. By continuing to control the on and off of the electrode, the microdroplet 201 containing the one cell 202 finally moves to a corresponding location of the second square electrode 2432. As a result, the liquid 200 containing a plurality of cells 202 forms a plurality of microdroplets 201 containing one cell 202 until the required amount of cells is selected, and a related cell experiment is performed at the corresponding location of the second square electrode 2432.

[0270] Specifically, in the above embodiment, the side length of the first square electrode 2431 is 50 μm to 2 mm, and the side length of the second square electrode 2432 is 1 / 5 to 1 / 2 the side length of the first square electrode 2431. The side length of the first hexagonal electrode 2441 is 50 μm to 2 mm, and the side length of the second hexagonal electrode 2442 is 1 / 5 to 1 / 2 the side length of the first hexagonal electrode 2441.

[0271] In some embodiments, referring to FIG. 34, the electrode layer 23 includes a plurality of first hexagonal electrodes 2441 arranged in an array, a plurality of second hexagonal electrodes 2442 arranged in an array, and a plurality of square electrodes 243 arranged in an array, which are connected in series.

[0272] Specifically, steps S1 to S6 in Fig. 34 show a specific application of the above embodiment. Liquid 200 enters the corresponding location of the first hexagonal electrode 2441, and by controlling the on and off of the electrode, the liquid 200 forms smaller volume droplets at the corresponding location of the second hexagonal electrode 2442. Then, by controlling the on and off of the electrode, the droplets at the corresponding location of the second hexagonal electrode 2442 form 20 petaliter-class microdroplets 201 at the corresponding location of the square electrode 243, and a related experiment of the microdroplets 201 at the corresponding location of the square electrode 243 is carried out.

[0273] Specifically, in the above embodiment, the side length of the square electrode 243 is 50 μm to 2 mm, the side length of the first hexagonal electrode 2441 is 50 μm to 2 mm, and the side length of the second hexagonal electrode 2442 is 1 / 5 to 1 / 2 of the side length of the first hexagonal electrode 2441.

[0274] In some embodiments, still referring to FIG. 28, the upper electrode plate 10 includes an upper cover 11, a conductive layer 12 and a first hydrophobic layer 13, which are arranged in sequence. The lower electrode plate 20 includes a second hydrophobic layer 21 and a dielectric layer 22. The second hydrophobic layer 21, the dielectric layer 22 and the electrode layer 23 are arranged in sequence. The first hydrophobic layer 13 and the second hydrophobic layer 21 are arranged opposite each other. A fluid passage layer 101 is formed between the first hydrophobic layer 13 and the second hydrophobic layer 21.

[0275] In some embodiments, the upper cover 11 has a thickness of 0.05 mm to 1.7 mm, the conductive layer 12 has a thickness of 10 nm to 500 nm, the dielectric layer 22 has a thickness of 50 nm to 1000 nm, the electrode layer 23 has a thickness of 10 nm to 1000 nm, the first hydrophobic layer 13 has a thickness of 10 nm to 200 nm, and the second hydrophobic layer 21 has a thickness of 10 nm to 200 nm.

[0276] In some embodiments, the material of the upper cover 11 can be glass-based. The material of the conductive layer 12 can be an ITO conductive layer. The material of the dielectric layer 22 can be an organic or inorganic insulating material. The material of the electrode layer 23 can be a metal or its oxide conductive material.

[0277] In some embodiments, the distance between the first hydrophobic layer 13 and the second hydrophobic layer 21 is 20 μm to 200 μm. Both the first hydrophobic layer 13 and the second hydrophobic layer 21 are made of hydrophobic materials, such as PTFE, fluorinated polyethylene, fluorocarbon wax, or other fluorine-containing polymers.

[0278] In some embodiments, the microfluidic chip includes a sample inlet hole (not shown) and a sample outlet hole (not shown). The sample inlet hole can inject liquid samples and media into the microfluidic chip, and the sample outlet hole can discharge liquid samples and media. Specifically, the sample inlet hole and the sample outlet hole can be provided on the upper electrode 10.

[0279] Based on the same concept, an embodiment of the present application also provides a micro-droplet generating method, as shown in Figure 35, including the following steps:

[0280] Step S11: providing the above-mentioned microfluidic chip.

[0281] Step S12: forming a plurality of suction points on the electrode plate under the microfluidic chip, which are used to suction liquid.

[0282] Step S13: Inject a liquid sample into the fluid passage layer of the microfluidic chip, and the liquid sample forms a micro-droplet at a location corresponding to the suction point.

[0283] In step S14, an attraction point is formed by an electrode that is turned on in the electrode layer of the microfluidic chip, and adjacent turned-on electrodes are separated by electrodes that are not turned on.

[0284] It should be noted that the micro-droplet generating method of the embodiment of the present application uses the above micro-fluidic chip to generate micro-droplets. The micro-fluidic chip 100 includes an upper electrode plate 10 and a lower electrode plate 20, and a fluid passage layer 101 is formed between the upper electrode plate 10 and the lower electrode plate 20. The lower electrode plate 20 forms a plurality of suction points, which are used to absorb the liquid 200. The lower electrode plate 20 includes an electrode layer 23. The electrode layer 23 includes a plurality of electrodes 24 arranged in an array having at least two different shapes. A liquid sample is injected into the fluid passage layer, and the liquid sample is attracted to the suction points, and the liquid sample leaves micro-droplets at the locations corresponding to the suction points by using the electrowetting principle. Using the above micro-droplet generating method, a high density of micro-droplets can be generated in a short time, the droplet generation time is greatly shortened, and the operation process is simple and convenient. Since no equipment such as a high-precision micro-pump is required, the system cost can be reduced. In addition, the method is scalable, and more micro-droplets and multiple sets of samples can be separated by expanding the size of the chip. In addition, the electrode layer includes multiple electrodes arranged in an array with at least two different shapes, so that by controlling the on and off of the electrodes, the large droplets can be formed by one type of multiple electrodes arranged in an array, and the related experiment of the microdroplets can be completed by another type of multiple electrodes arranged in an array, thereby avoiding the transfer of liquid samples.

[0285] In some embodiments, a method for generating microdroplets is as follows: A medium is injected into a fluid passage layer of a microfluidic chip, and the fluid passage layer is filled with the medium. Specifically, the medium can be air, silicone oil, or mineral oil.

[0286] Next, a liquid sample is injected into the fluid passage layer of the microfluidic chip, and the liquid sample is enveloped in the medium, forming a microdroplet at a location corresponding to the suction point.

[0287] Sixth embodiment As shown in Figures 36 to 42, the detailed structure of a micro-droplet generating system and a micro-droplet generating method according to the sixth embodiment of the present application are specifically described.

[0288] Referring to Figure 36, the present application presents a method for rapid generation of microdroplets, which involves the following steps:

[0289] Step S71: Provide a microfluidic chip. The microfluidic chip comprises an upper electrode plate 10 and a lower electrode plate 20. A fluid passage layer is formed between the upper electrode plate and the lower electrode plate. The lower electrode plate 20 includes an electrode layer 23. The electrode layer 23 includes a plurality of electrodes 24 arranged in an array.

[0290] In step S72, a plurality of attraction points are formed on the lower electrode plate 20. The attraction points are used to attract liquid. The attraction points are formed from the electrodes 241 that are turned on in the electrode layer 23. Adjacent electrodes 241 that are turned on are spaced apart by electrodes 242 that are not turned on.

[0291] In step S73, a liquid sample is injected into the fluid passage layer 101, and the electrodes 24 are turned on and off to cause the liquid sample to flow through the fluid passage layer 101 at a position corresponding to the suction point. 1 Microdroplets are formed.

[0292] Step S74: Also, by controlling the on and off of the electrode 24, the n 1 Of the n microdroplets, all of them are attracted to the suction point. 2 Microdroplets are formed.

[0293] Step S75: Further, the electrode 24 is turned on and off to control the n 2 Of the n microdroplets, all the microdroplets form n3 microdroplets at the suction point. In step S76, the electrodes are repeatedly turned on and off until the desired number of microdroplets are formed. Of these, the above n 1 , n2 , n 3 is a positive integer equal to or greater than 2.

[0294] It should be noted that in the high-speed microdroplet generation method described in the embodiment of the present application, a liquid sample is poured into the fluid passage layer 101, and the liquid sample fills the fluid passage layer 101. The liquid sample flows in the fluid passage layer 101 and forms microdroplets at the locations corresponding to the attraction points. Specifically, by controlling the on and off of the electrodes 24 of the electrode layer 23, the electrowetting principle (when liquid exists in the electrode, the wettability of the solid-electrolyte interface at the location corresponding to the electrode can be changed by applying an electric field to the electrode. This also changes the contact angle between the droplet and the electrode interface. Due to the electric field difference between the electrodes in the droplet area, when the contact angles are different, a lateral propulsion force is generated and the droplet moves laterally on the electrode substrate) is utilized, and the liquid sample is attracted by the turned-on electrodes to form multiple microdroplets at the locations corresponding to the multiple electrodes turned on in the fluid passage layer. The attraction points are formed by the electrodes 241 turned on in the electrode layer 23, and the adjacent turned-on electrodes 241 are separated by the electrodes 242 that are not turned on. Furthermore, the movement of the microdroplets can be controlled by controlling the on and off of the electrodes. The specific method of forming the microdroplets from the liquid sample is as follows: By controlling the on and off of the electrodes 24, the liquid sample moves n 1 Next, the n microdroplets are formed by controlling the on and off of the electrode 24. 1 Of the n microdroplets, all of them are attracted to the n 2 Furthermore, by controlling the on / off of the electrode 24, the n microdroplets formed are 2 Of the n microdroplets, all of them are attracted to the n 3 The electrodes 24 are turned on and off repeatedly until the desired number of microdroplets are formed, and all of the formed microdroplets continue to form multiple microdroplets. 1 , n 2 , n 3is a positive integer equal to or greater than 2. Specifically, n 1 , n 2 , n 3 n can also be 2, 3, 4, 5, 6, 7, 8, 9, or 10. 1 , n 2 , n 3 The value of may be the same or different. That is, there is no correlation between the number of microdroplets formed in two times, and the more microdroplets formed in one time, the better the efficiency of generating microdroplets. For example, the liquid sample forms 10 microdroplets at a position corresponding to the suction point, and by controlling the on and off of the electrode 24, all of the formed 10 microdroplets form 10 microdroplets again at the suction point (8 or 11 are also possible, and the number of microdroplets is specifically formed according to needs) at the suction point. By continuously controlling the on and off of the electrode 24, all of the formed 10 microdroplets form 10 microdroplets at the suction point. By repeatedly turning on and off the electrode 24, 10 microdroplets are finally formed. n Using the high-speed microdroplet generation method of the present invention, a large amount of microdroplets can be formed in a short time, and the required number of microdroplets can be generated at high speed, improving the efficiency and flow rate of microdroplets. The high-speed microdroplet generation method of the present invention is advantageous in experiments that require a large number of droplets (digital PCR, digital Elisa, generating single cells).

[0295] Specifically, adjacent turned-on electrodes 241 are separated by non-turned-on electrodes 242. Optimally, adjacent turned-on electrodes 241 are separated by at least two non-turned-on electrodes 242.

[0296] In some embodiments, a liquid sample is injected into the fluid passage layer 101, and by controlling the electrodes 24 to be on and off, the liquid sample forms two micro-droplets at the positions corresponding to the attraction points.

[0297] Then, by controlling the electrode 24 to be turned on and off, every two micro-droplets formed form two micro-droplets at the position of the attraction point.

[0298] Subsequently, by controlling the on and off of the electrode 24, every two micro-droplets formed form two micro-droplets at the position of the attraction point.

[0299] In the above embodiment, referring to FIG. 37, the electrode 24 is in the shape of a square, and the liquid 200 first forms two microdroplets after the liquid sample moves by controlling the electrode 24 on and off, and then, by controlling the electrode 24 on and off, every two microdroplets formed form two more microdroplets. At this point, four microdroplets are formed in total. Then, the electrode 24 is controlled on and off again, and every microdroplet formed forms two more microdroplets, and eight microdroplets are formed in total at this point. Then, the electrode 24 is controlled on and off again, and every microdroplet formed forms two more microdroplets, and sixteen microdroplets are formed in total at this point. Repeating in this manner, two microdroplets are finally formed. n Microdroplets are formed.

[0300] In some embodiments, a liquid sample is injected into the fluid passage layer 101, and by controlling the electrodes 24 to be on and off, the liquid sample forms three micro-droplets at the locations corresponding to the attraction points.

[0301] Then, by controlling the electrode 24 to be turned on and off, all three micro-droplets formed form three micro-droplets at the location of the attraction point.

[0302] Subsequently, by controlling the on and off of the electrode 24, all three micro-droplets formed form three micro-droplets at the position of the attraction point.

[0303] The electrodes 24 are repeatedly turned on and off to form a desired number of microdroplets.

[0304] In the above embodiment, by controlling the on and off of the electrode 24, after the liquid sample moves, first three microdroplets are formed, and then by controlling the on and off of the electrode 24, all of the three microdroplets formed once again form three microdroplets, forming a total of nine microdroplets at this point. Then, by controlling the on and off of the electrode 24 once again, all of the formed microdroplets form three microdroplets once again, forming a total of 27 microdroplets at this point. Then, by controlling the on and off of the electrode 24 once again, all of the formed microdroplets form three microdroplets once again, forming a total of 81 microdroplets at this point. Repeating in this manner, finally, three n Microdroplets are formed.

[0305] In some embodiments, a liquid sample is injected into the fluid passage layer 101, and by controlling the electrodes 24 to be on and off, the liquid sample forms four micro-droplets at the locations corresponding to the attraction points.

[0306] Then, by controlling the electrodes 24 to be turned on and off, all four micro-droplets formed form four micro-droplets at the location of the attraction point.

[0307] Subsequently, by controlling the on and off of the electrode 24, all the four micro-droplets formed form four micro-droplets at the position of the attraction point.

[0308] The electrodes 24 are repeatedly turned on and off to form a desired number of microdroplets.

[0309] In the above embodiment, by controlling the on and off of the electrodes 24, after the liquid sample has moved, first four microdroplets are formed, and then by controlling the on and off of the electrodes 24, all of the four microdroplets formed once again form four microdroplets, forming a total of 16 microdroplets at this point. Then, by controlling the on and off of the electrodes 24 once again, all of the microdroplets formed once again form four microdroplets, forming a total of 64 microdroplets at this point. Then, by controlling the on and off of the electrodes 24 once again, all of the microdroplets formed once again form four microdroplets, forming a total of 256 microdroplets at this point. This process is repeated until finally 4 n Microdroplets are formed.

[0310] In some embodiments, the electrodes 24 are square or hexagonal in shape. It should be understood that hexagonal electrodes allow droplet breakup in six directions, which is advantageous over the four directions of a square. In addition to squares and hexagons, the electrodes may be any shape or combination of shapes.

[0311] In some embodiments, the side length of electrode 24 is between 50 μm and 2 mm.

[0312] By adjusting the size of the electrodes and the gap distance between the electrodes, the volume of the microdroplets can be precisely tuned. Furthermore, by controlling the position and number of electrodes that are turned on, the position and number of microdroplets formed can be controlled, which means the density at which the microdroplets form can be precisely controlled.

[0313] FIG. 38 shows the actual experimental process of liquid movement and generating microdroplets in the sixth embodiment of the present application. Specifically, in the figure, the electrode 24 is in the shape of a square, and the liquid 200 first forms two microdroplets after the liquid sample moves by controlling the on and off of the electrode 24, and then, by controlling the on and off of the electrode 24, every two microdroplets formed form two microdroplets again. At this point, a total of four microdroplets are formed. Then, the electrode 24 is controlled on and off again, and every microdroplet formed forms two microdroplets again, and a total of eight microdroplets are formed at this point. Then, the electrode 24 is controlled on and off again, and every microdroplet formed forms two microdroplets again, and a total of 16 microdroplets are formed at this point. Then, the electrode 24 is controlled on and off again, and every microdroplet formed forms two microdroplets again, and a total of 32 microdroplets are formed at this point.

[0314] FIG. 39 shows the experimental process of single cell separation in the sixth embodiment of the present application, in which the liquid moves and generates microdroplets, using the method 1. In the figure, the electrode 24 is square-shaped, and the liquid 200 controls the on and off of the electrode 24 to first form 16 microdroplets after the liquid sample moves, and then controls the on and off of the electrode 24 so that each of the 16 microdroplets forms two more microdroplets. At this point, a total of 32 microdroplets are formed. This is the process of the single cell separation experiment in which the liquid sample moves and generates microdroplets in the sixth embodiment. The difference from FIG. 38 is that this method generates droplets containing single cells.

[0315] In some embodiments, referring to Figure 40, in which the electrodes 24 are square shaped, the liquid 200 first forms three microdroplets after the liquid sample moves by controlling the electrodes 24 on and off, and then every two microdroplets formed form three microdroplets again by controlling the electrodes 24 on and off, forming a total of nine microdroplets at this point, and then again by controlling the electrodes 24 on and off, forming every microdroplet formed form two microdroplets again, forming a total of 18 microdroplets at this point.

[0316] In the above embodiment, referring to FIG. 41, the electrode 24 in the figure is hexagonal, and the liquid 200 first forms two microdroplets after the liquid sample moves by controlling the electrode 24 on and off, and then every two microdroplets formed form two more microdroplets by controlling the electrode 24 on and off. At this point, a total of four microdroplets are formed. Then, the electrode 24 is controlled on and off again, and every microdroplet formed forms two more microdroplets, and a total of eight microdroplets are formed at this point. Then, the electrode 24 is controlled on and off again, and every microdroplet formed forms two more microdroplets, and a total of 16 microdroplets 201 are formed at this point.

[0317] In some embodiments, referring to Figure 42, in which the electrodes 24 are hexagonal in shape, the liquid 200 first forms three micro-droplets after the liquid sample moves by controlling the electrodes 24 on and off, and then by controlling the electrodes 24 on and off, every three micro-droplets formed form three more micro-droplets, forming a total of nine micro-droplets at this point, and then by controlling the electrodes 24 on and off again, every micro-droplet formed forms two more micro-droplets, forming a total of 18 micro-droplets at this point.

[0318] The structure of the microfluidic chip of the sixth embodiment of the present application is the same as that of the fifth embodiment. As shown in FIG. 28, in the sixth embodiment, the upper electrode plate 10 includes an upper cover 11, a conductive layer 12 and a first hydrophobic layer 13, which are sequentially arranged. The lower electrode plate 20 includes a second hydrophobic layer 21 and a dielectric layer 22. The second hydrophobic layer 21, the dielectric layer 22 and the electrode layer 23 are sequentially arranged in a stacked manner. The first hydrophobic layer 13 and the second hydrophobic layer 21 are arranged opposite to each other. A fluid passage layer 101 is formed between the first hydrophobic layer 13 and the second hydrophobic layer 21.

[0319] In some embodiments, the upper cover 11 has a thickness of 0.05 mm to 1.7 mm, the conductive layer 12 has a thickness of 10 nm to 500 nm, the dielectric layer 22 has a thickness of 50 nm to 1000 nm, the electrode layer 23 has a thickness of 10 nm to 1000 nm, the first hydrophobic layer 13 has a thickness of 10 nm to 200 nm, and the second hydrophobic layer 21 has a thickness of 10 nm to 200 nm.

[0320] In some embodiments, the material of the upper cover 11 can be glass-based. The material of the conductive layer 12 can be an ITO conductive layer. The material of the dielectric layer 22 can be an organic or inorganic insulating material. The material of the electrode layer 23 can be a metal or its oxide conductive material.

[0321] In some embodiments, the distance between the first hydrophobic layer 13 and the second hydrophobic layer 21 is 5 μm to 600 μm. Both the first hydrophobic layer 13 and the second hydrophobic layer 21 are made of hydrophobic materials, such as PTFE, fluorinated polyethylene, fluorocarbon wax, or other fluorine-containing polymers.

[0322] In some embodiments, a method for generating microdroplets is as follows.

[0323] A medium is injected into the fluid passage layer of the microfluidic chip, so that the fluid passage layer 101 is filled with the medium. Next, a liquid sample is injected into the fluid passage layer of the microfluidic chip, so that the liquid sample is surrounded by the medium, and the liquid sample forms a microdroplet at a location corresponding to its suction point.

[0324] Specifically, the medium may be air, silicone oil, mineral oil, or the like.

[0325] In some embodiments, the microfluidic chip includes a sample inlet hole (not shown) and a sample outlet hole (not shown). The sample inlet hole can inject liquid samples and media into the microfluidic chip, and the sample outlet hole can discharge liquid samples and media. Specifically, the sample inlet hole and the sample outlet hole can be provided on the upper electrode 10.

[0326] Generally speaking, based on the first to sixth embodiments of the present application, the present application puts forward a method for generating micro-droplets, which includes the following steps:

[0327] Step S1: Provide a microfluidic chip 100. The microfluidic chip 100 comprises an upper electrode plate 10 and a lower electrode plate 20. A fluid passage layer 101 is formed between the upper electrode plate 10 and the lower electrode plate 20.

[0328] In step S2, at least one of the upper plate 10 and the lower plate 20 forms a plurality of suction points, which are used to suck in the liquid 200.

[0329] In step S3, the liquid 200 is poured into the fluid passage layer 101.

[0330] In step S4, the liquid 200 is driven to flow in the fluid passage layer 101 to form micro-droplets 201 at multiple suction points of the microfluidic chip 100.

[0331] In the present application, the above-mentioned microdroplet generating method can generate a large amount of microdroplets 201 in a short time, greatly shorten the droplet generating time, and the operation process is simple and convenient. Since equipment such as a high-precision micropump is not required, the system cost can be reduced. In addition, the method is scalable, and more microdroplets and multiple sets of samples can be separated by expanding the size of the microfluidic chip 100. In addition, the volume and density of the formed microdroplets can be precisely adjusted by adjusting the gap between the upper and lower plates, the number of suction points, the area size and the position mode. Therefore, the present invention provides a microdroplet generating method and a microdroplet generating system that can rapidly form high-density microdroplets and precisely control the volume and density of the formed high-density microdroplets.

[0332] Based on the above description of the disclosed embodiments, a person skilled in the art can realize or use the present application. It is easy for a person skilled in the art to apply the present application by referring to the above embodiments. The general principles defined herein may be realized in other embodiments within the content or scope of the present application. Thus, the present application should not be limited to the embodiments shown herein, but should be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A micro-droplet generating system, comprising: a micro-fluidic chip; and a droplet driving unit connected to the micro-fluidic chip, the micro-fluidic chip comprising an upper electrode plate and a lower electrode plate, a fluid passage layer is formed between the upper electrode plate and the lower electrode plate, and an attraction point is used for attracting liquid, the upper electrode plate comprises an upper cover, a conductive layer and a first hydrophobic layer, which are stacked in order, the lower electrode plate comprises a second hydrophobic layer, a dielectric layer, an electrode layer and a substrate, which are stacked in order, a hydrophilic point array is formed on the first hydrophobic layer of the upper substrate away from the conductive layer, the hydrophilic points of the hydrophilic point array are the attraction points, and the attraction points are formed by electrodes of the electrode layer that are turned on, and the droplet driving unit is used to drive a flow in the fluid passage layer by a liquid injected into the fluid passage layer, thereby forming micro-droplets at the positions of the attraction points.

2. The micro-droplet generating system according to claim 1, characterized in that the first hydrophobic layer and the second hydrophobic layer are arranged opposite each other, the fluid passage layer is formed between the first hydrophobic layer and the second hydrophobic layer, and the electrode layer is composed of a plurality of electrodes arranged in an array.

3. The micro-droplet generation system according to claim 2, characterized in that the attraction points are formed from the electrodes of the electrode layer that are turned on, and adjacent turned-on electrodes are separated by turned-off electrodes.

4. 3. The micro-droplet generating system according to claim 2, wherein adjacent hydrophilic points in the hydrophilic point array are separated by the attraction points.

5. The micro-droplet generating system according to claim 2 , wherein the shape of the electrodes of the electrode layer is a hexagon or a square.

6. The micro-droplet generation system according to claim 2 , wherein the electrode layer includes a plurality of square electrodes arranged in an array and a plurality of hexagonal electrodes arranged in an array.

7. The microdroplet generation system of claim 6, characterized in that the electrode layer includes a plurality of hexagonal electrodes arranged in an array and a plurality of square electrodes arranged in an array on both sides of the plurality of hexagonal electrodes arranged in an array.

8. The microdroplet generation system described in claim 6, characterized in that the electrode layer includes a plurality of square electrodes arranged in an array and a plurality of hexagonal electrodes arranged in an array on both sides of the plurality of square electrodes arranged in an array.

9. 9. The micro-droplet generating system according to claim 7 or 8, wherein the side length of the hexagonal electrode is 50 μm to 2 mm, and the side length of the square electrode is 50 μm to 2 mm.

10. The microdroplet generation system of claim 6, characterized in that the electrode layer includes a plurality of first square electrodes arranged in an array, a plurality of first hexagonal electrodes arranged in an array, a plurality of second hexagonal electrodes arranged in an array, and a plurality of second square electrodes arranged in an array, which are connected in sequence.

11. The microdroplet generation system of claim 6, characterized in that the electrode layer includes a plurality of first hexagonal electrodes arranged in an array, a plurality of second hexagonal electrodes arranged in an array, and a plurality of square electrodes arranged in an array, which are connected in sequence.

12. A microdroplet generation system as described in claim 10 or 11, characterized in that the side length of the first square electrode or the square electrode is 50 μm to 2 mm, the side length of the second square electrode is 1 / 5 to 1 / 2 of the side length of the first square electrode, the side length of the first hexagonal electrode is 50 μm to 2 mm, and the side length of the second hexagonal electrode is 1 / 5 to 1 / 2 of the side length of the first hexagonal electrode.

13. The micro-droplet generating system of any one of claims 2 to 5, characterized in that the droplet driving unit includes an electrode driving unit connected to the electrode layer and used to control the on and off of the electrodes of the electrode layer, thereby forming micro-droplets at the suction point by controlling the flow of the liquid injected into the fluid passage layer in the fluid passage layer.

14. The micro-droplet generating system according to any one of claims 2 to 5, characterized in that a liquid injection hole is provided at the center of the micro-fluidic chip, which is used to inject liquid into the fluid passage layer; the micro-fluidic chip is provided with a plurality of liquid discharge holes, which are used to discharge excess liquid from the micro-fluidic chip; the droplet driving unit includes a rotation driving unit, which is used to drive the rotation of the micro-fluidic chip, so that the liquid injected into the fluid passage layer forms micro-droplets at the suction point by a spin coating method.

15. The micro-droplet generating system according to claim 14 , wherein when the rotation drive unit is driven, the rotation speed of the micro-fluidic chip is greater than 0 rpm and equal to or less than 1000 rpm.

16. The micro-droplet generating system according to claim 14, characterized in that the electrode is hexagonal in shape, the side length is 50 μm to 2 mm, and the distance between the first hydrophobic layer and the second hydrophobic layer is 5 μm to 600 μm.

17. 6. The micro-droplet generating system according to claim 2, wherein the micro-fluidic chip is provided with a first sample inlet hole and a first sample outlet hole, the first sample inlet hole and the first sample outlet hole being arranged on a first diagonal of the micro-fluidic chip, the droplet driving unit includes a first micro-pump and a third micro-pump, the first micro-pump is connected to the first sample inlet hole and is used to inject liquid into the fluid passage layer, thereby filling the fluid passage layer with the liquid, and the third micro-pump is connected to the first sample outlet hole and is used to pump the liquid or gas flowing out from the first sample outlet hole, thereby forming a micro-droplet at the suction point.

18. The micro-droplet generating system of claim 17, characterized in that the micro-fluidic chip further has a second sample inlet hole and a second sample outlet hole, the second sample inlet hole and the second sample outlet hole are arranged on a second diagonal of the micro-fluidic chip, the droplet driving unit further includes a second micro-pump and a fourth micro-pump, the second micro-pump is connected to the second sample inlet hole and is used to inject a medium into the fluid passage layer, and the fourth micro-pump is connected to the second sample outlet hole and is used to pump up excess liquid or medium flowing out from the second sample outlet hole, thereby causing the medium to be enveloped in the micro-droplet formed at the suction point.

19. The micro-droplet generating system according to claim 17, wherein the upper cover has a thickness of 0.05 mm to 1.7 mm, the substrate has a thickness of 0.05 mm to 1.7 mm, the conductive layer has a thickness of 10 nm to 500 nm, the dielectric layer has a thickness of 50 nm to 1000 nm, the electrode layer has a thickness of 10 nm to 1000 nm, the first hydrophobic layer has a thickness of 10 nm to 200 nm, and the second hydrophobic layer has a thickness of 10 nm to 200 nm.

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