A microfluidic chip based on droplet splitting and fusion and a manufacturing method thereof

Abstract

The application discloses a micro-fluidic chip based on liquid drop splitting and fusion, through a T-shaped structure of an input end and a multi-stage Y-shaped structure of an output end, liquid concentration can be accurately controlled in the dimensions of space and time, samples and diluent are mixed by using the principle of liquid drop splitting and fusion, the size of the liquid drop is controllable, the concentration of the generated solution is simple and controllable to generate a multi-phase micro-fluid to produce a small liquid drop with a controllable concentration gradient at the end. The micro-fluidic chip does not need to control the flow rate by using an accurate micro-fluid pump, only a common syringe is used to realize a specific concentration gradient, and the system cost is greatly reduced. The application further provides a manufacturing method of the micro-fluidic chip.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of microfluidic devices, and particularly relates to a microfluidic chip based on droplet splitting and fusion and a manufacturing method thereof. BACKGROUND

[0002] Microfluidic technology is an effective technology platform that can provide stable gradients for cell behavior research. Recently, it has been proposed that a universal gradient generation device can generate gradients from only two starting solutions. A multistage solution distributor is provided in the microchannel to limit diffusion mixing, so that gradients of arbitrary shape can be generated. However, a complex calculation is involved to determine the relative positions of the gradient distribution solution splitters. In addition, all the above-mentioned gradient generation devices are designed to generate gradients transverse to the main flow. Chemical gradients can also be generated along the main flow in a microfluidic device. Chemical gradients are generated by diffusion of sample molecules. However, such gradient distribution has limited controllability.

[0003] In the research and application of microfluidic systems, a key and indispensable link is the fabrication of microfluidic molds. At present, the commonly used mold fabrication method at home and abroad is based on traditional photolithography technology. "Photolithography" refers to the process of transferring patterns on a mask plate to a silicon wafer coated with photoresist, and then going through post-baking, developing, hard-baking, etching, and other processes to successfully fabricate microchannel structures on a silicon or glass substrate. The entire photolithography process must be completed by specially trained personnel in a standard clean laboratory environment, and the quality and precision requirements of the mask used in photolithography are relatively high (usually a standard chromium mask is required).

[0004] At present, although the prior art can generate concentration gradients at the microscale, it has limitations. Most concentration gradient generation techniques cannot accurately control the distribution of the concentration gradient as required, and only rely on changing the concentration of the injected fluid to change the concentration gradient, generating a single gradient. In addition, many concentration gradient generation devices have complex structures and have high requirements for the manufacturing process of the device. At the same time, after the liquid enters the channel network from the concentration gradient generation device, part of the liquid may be trapped in the corners of the channel, thereby interfering with the normal flow of the fluid, such as the most commonly used Christmas tree structure. And if you want to use the Christmas tree structure to generate an ultra-long concentration gradient, a large number of microchannel structures need to be added to form a large area and complex interlaced microchannel network.

[0005] Photolithography demands stringent substrate surface quality, is complex, and requires expensive exposure and etching equipment, resulting in high costs, typically around one million yuan. Mask utilization rates are low, and the entire photolithography process must be completed in a standard cleanroom environment by specially trained personnel strictly following the fabrication procedures. The complexity, high cost, and long cycle time of photolithography are major bottlenecks restricting the research and development of microfluidic technology in non-clean environments. Traditional microfluidic chip fabrication techniques hinder the rapid fabrication and preparation of microfluidic chip microstructures. Summary of the Invention

[0006] To address the technical problems existing in the background art, this invention proposes a microfluidic chip based on droplet splitting and fusion and its fabrication method.

[0007] The present invention proposes a microfluidic chip based on droplet splitting and fusion, comprising: an input section and an output section;

[0008] The input section includes a first input terminal, a second input terminal, a dispersed phase input terminal, and two T-shaped input structures. The longitudinal ends of the two T-shaped structures are connected to the dispersed phase input terminal, and the first input terminal and the second input terminal are respectively connected to the first transverse ends of the two T-shaped input structures.

[0009] The output section includes a primary output structure and at least one secondary output structure. The primary output structure includes two Y-shaped structures, which are arranged in parallel and whose main ends are respectively connected to the second lateral ends of two T-shaped input structures. The adjacent branch ends of the two Y-shaped structures form a primary mixing chamber that is interconnected with each other. The primary mixing chamber has a mixing output port.

[0010] The secondary output structure includes two end Y-shaped structures and at least one middle Y-shaped structure located between the two end Y-shaped structures. The end Y-shaped structures and the middle Y-shaped structures are arranged in parallel. The main end of the end Y-shaped structure is connected to the outer branch end of the upper-level Y-shaped structure. The main end of the middle Y-shaped structure is connected to the mixing output port of the primary mixing chamber. Secondary mixing chambers that are connected to each other are formed at the branch ends of adjacent Y-shaped structures. The secondary mixing chambers have mixing output ports.

[0011] The hybrid output port of the terminal secondary output structure and the outer branch end of the terminal Y-shaped structure serve as droplet output ends.

[0012] Preferably, the output section includes multiple secondary output structures, the number of middle Y-shaped structures of the multiple secondary output structures gradually increases in the direction away from the primary output structure, the main end of each end Y-shaped structure is connected to the outer branch end of the previous end Y-shaped structure, and the main end of the middle Y-shaped structure is connected to the mixing output port of the previous mixing chamber.

[0013] Preferably, the Y-shaped structures of the output section have the same dimensions.

[0014] In the present application, the microfluidic chip based on droplet splitting and fusion can accurately control the liquid concentration in the dimensions of space and time by the input T-shaped structure and the output multi-stage Y-shaped structure design, mix the sample and the diluent by the principle of droplet splitting and fusion, the droplet size is controllable, the solution concentration calculation is simple and controllable, and the multi-phase micro fluid generates small droplets with controllable concentration gradient at the end. The microfluidic chip of the present application does not need to accurately control the flow rate of the micro pump at the input end, and only a common syringe can realize a specific concentration gradient, which greatly reduces the system cost.

[0015] In addition, super multi-component concentration gradient can be realized at the same time, so that biological and chemical reactions can be carried out at different drug concentrations. And the concentration gradient fluid is divided into droplets, each droplet can be used as a separate micro reactor, which not only reduces cross contamination, but also reduces the volume of the chip.

[0016] The present application also provides a manufacturing method of the microfluidic chip based on droplet splitting and fusion, comprising the following steps:

[0017] S1: mask plate manufacturing; creating a chip mask pattern, and printing a transparent mask sheet according to the chip mask pattern;

[0018] S2: male mold manufacturing; covering two layers of photosensitive dry film on the surface of the glass slide to form a glass slide with a photosensitive dry film layer, using a UV ultraviolet lamp to expose the glass slide to the transparent mask plate, so that the chip pattern on the mask is transferred to the photosensitive dry film layer by the principle of exposure imaging, then the exposed glass slide is baked and developed with developer, and finally heated and cured to obtain the photosensitive dry film with the chip mask pattern as the male mold;

[0019] S3: microfluidic chip manufacturing: stirring the prepolymer and curing agent of PDMS to make the gas bubbles in the mixed liquid uniformly distributed, vacuumizing the mixed liquid by a vacuum dryer until the gas bubbles in the mixed liquid disappear; then pouring the mixed liquid into the male mold, and placing it in an oven for curing; cutting the chip mask pattern to demold, and punching holes at the inlet and outlet to form a chip embryo; finally, performing oxygen plasma treatment on the chip embryo, covering the photosensitive dry film side of the glass slide with a cover glass, and bonding them by lock reaction to obtain a microfluidic chip.

[0020] Preferably, in S3, the ratio of the prepolymer and the curing agent is 10:1.

[0021] Preferably, in S3, the mixed liquid is poured into the male mold, specifically, the edge of the male mold is wrapped with tin paper first, then placed in a culture dish, and then the prepared mixed liquid is slowly poured in to ensure that the PDMS is uniformly distributed in the male mold and the liquid level is level.

[0022] Preferably, in S3, the mixed solution is poured into the positive mold and then vacuumized again.

[0023] Preferably, in S3, the wafer embryo is treated by oxygen plasma, specifically, the cover glass and the glass slide are put into a plasma cleaner, the radio frequency power is 45 W, and the oxygen plasma treatment time is 3 min.

[0024] In the present application, the preparation method of the microfluidic chip based on droplet splitting and fusion is simple, low in cost, and can provide an economical scientific research experimental platform for the application of microfluidic chips. BRIEF DESCRIPTION OF DRAWINGS

[0025] Figure 1 FIG. 1 is a structural schematic diagram of an embodiment of a microfluidic chip based on droplet splitting and fusion according to the present application.

[0026] Figure 2 FIG. 2 is a structural schematic diagram of an input part of an embodiment of a microfluidic chip based on droplet splitting and fusion according to the present application.

[0027] Figure 3 FIG. 3 is a structural schematic diagram of a first-stage output structure of an embodiment of a microfluidic chip based on droplet splitting and fusion according to the present application.

[0028] Figure 4 FIG. 4 is a structural schematic diagram of an output end of an embodiment of a microfluidic chip based on droplet splitting and fusion according to the present application.

[0029] Figure 5 FIG. 5 is a flowchart of an embodiment of a preparation method of a microfluidic chip based on droplet splitting and fusion according to the present application.

[0030] Figure 6 FIG. 6 is a sample diagram of an embodiment of a microfluidic chip based on droplet splitting and fusion according to the present application. DETAILED DESCRIPTION

[0031] As shown in FIG. 1, FIG. 2, FIG. 3, FIG. 4, and FIG. 5, the present application provides a microfluidic chip based on droplet splitting and fusion. Figures 1 to 6 As shown in FIG. 1, FIG. 2, FIG. 3, FIG. 4, and FIG. 5, the present application provides a microfluidic chip based on droplet splitting and fusion. Figure 1 As shown in FIG. 1, FIG. 2, FIG. 3, FIG. 4, and FIG. 5, the present application provides a microfluidic chip based on droplet splitting and fusion. Figure 2 As shown in FIG. 1, FIG. 2, FIG. 3, FIG. 4, and FIG. 5, the present application provides a microfluidic chip based on droplet splitting and fusion. Figure 3 As shown in FIG. 1, FIG. 2, FIG. 3, FIG. 4, and FIG. 5, the present application provides a microfluidic chip based on droplet splitting and fusion. Figure 4A structural schematic diagram of an output end of an embodiment of a microfluidic chip based on droplet splitting and fusion according to the present application, Figure 5 A flow chart of an embodiment of a manufacturing method of a microfluidic chip based on droplet splitting and fusion according to the present application, Figure 6 A sample diagram of an embodiment of a microfluidic chip based on droplet splitting and fusion according to the present application.

[0032] Referring to Figures 1 to 4 The microfluidic chip based on droplet splitting and fusion according to the present application comprises an input part and an output part.

[0033] The input part comprises a first input end 1, a second input end 2, a dispersed phase input end 3, and two T-shaped input structures, the longitudinal ends of the two T-shaped structures being connected to the dispersed phase input end 3, and the first input end 1 and the second input end 2 being connected to the first transverse ends of the two T-shaped input structures respectively.

[0034] The output part comprises a primary output structure and at least one secondary output structure, the primary output structure comprising two Y-shaped structures, the two Y-shaped structures being arranged in parallel and the main road ends of the two Y-shaped structures being connected to the second transverse ends of the two T-shaped input structures respectively, adjacent branch ends of the two Y-shaped structures forming a primary mixing chamber 4 which communicates with each other and has a mixing output port.

[0035] The secondary output structure comprises two end Y-shaped structures and at least one middle Y-shaped structure between the two end Y-shaped structures, the end Y-shaped structures and the middle Y-shaped structure being arranged in parallel, the main road end of the end Y-shaped structure communicating with the outer branch end of the Y-shaped structure of the previous stage, the main road end of the middle Y-shaped structure communicating with the mixing output port of the primary mixing chamber 4, the branch ends of adjacent Y-shaped structures forming secondary mixing chambers 5 which communicate with each other and have mixing output ports.

[0036] The mixing output ports of the end secondary output structures and the outer branch ends of the end Y-shaped structures serve as droplet output ends 6.

[0037] An auxiliary channel is arranged in the middle of the two main channels, the main channel inlets are used to input drug reagents and auxiliary functional reagents or diluents of the drug reagents, and the auxiliary channel inlets in the middle are used to input silicone oil. The auxiliary channel outlets communicate with the two main channels transversely to form two T-shaped structures. Two groups of droplets generated by the T-shaped structures are split into two groups of small droplets by the inverted Y-shaped structure, and then merged into the diffusion channel by the Y-shaped structure for fusion. Micro droplets are generated by using multi-phase micro fluid, the droplet size is controlled by adjusting the flow rate of the injection pump, and each droplet finally generated is regarded as a micro reactor.

[0038] The input part of the embodiment is provided with two T-shaped structures, two groups of liquid drops are generated at the same time, the liquid drops are split through the inverted Y-shaped structure design, and the liquid drops are fused by using the Y-shaped structure and the expansion structure. In the embodiment, the concentration gradient is not relied on diffusion mixing, but the diluent and the sample are divided into two groups of liquid drops A and B, the liquid drops are split into two groups when flowing through the bifurcation structure, and the two groups are A11, A22 and B11, B12. A11 meets B11 in the Y-shaped structure, and is mixed in the expansion structure to generate a liquid drop string as the first concentration liquid drop. The first concentration liquid drop is split into two groups again when flowing through the bifurcation structure, one group meets A22 in the Y-shaped structure and is mixed to be diluted into the second concentration liquid drop with a concentration smaller than the first concentration, and the other group meets B12 in the Y-shaped structure and is mixed to be diluted into the third concentration liquid drop with a concentration larger than the first concentration. The chemical gradient distribution along the main stream can control the step-by-step and convective distribution of the sample solution. The key part of the step-by-step distribution is the microchannel to the Y-shaped structure. By increasing the number of split mixing structures, a complex and controllable gradient distribution can be generated.

[0039] In the embodiment, the microfluidic chip based on liquid drop splitting and fusion is designed through the input end T-shaped structure and the output end multi-stage Y-shaped structure, so that the liquid concentration can be accurately controlled in the spatial and temporal dimensions. The sample and the diluent are mixed by using the principle of liquid drop splitting and fusion, the liquid drop size is controllable, the solution concentration calculation is simple and controllable, and the multi-phase micro fluid generating micro liquid drops with controllable concentration gradient is formed at the end. The microfluidic chip of the embodiment does not need to accurately control the flow rate of the micro flow pump at the input end, and only a common syringe is needed to realize a specific concentration gradient, so that the system cost is greatly reduced.

[0040] In the specific design of the output part, the output part includes a plurality of secondary output structures, the number of Y-shaped structures in the middle of the plurality of secondary output structures gradually increases away from the primary output structure, the main road end of each end Y-shaped structure is in communication with the outer side branch end of the end Y-shaped structure of the previous stage, and the main road end of the middle Y-shaped structure is in communication with the mixed output port of the mixing chamber 4 of the previous stage. According to the need, a plurality of output structures can be arranged to make the sample and the diluent perform gradient distribution mixing, and finally obtain liquid drops with gradually increasing or decreasing concentration from one end Y-shaped branch to the other end at the end. By using the multi-stage structure arrangement, the number of stages can be increased or reduced according to the detection requirements to configure the required concentration gradient.

[0041] In actual design, the Y-shaped structures of the output part are of the same size.

[0042] The embodiment also provides a manufacturing method of the microfluidic chip based on liquid drop splitting and fusion.

[0043] S1: mask plate manufacturing; creating a chip mask pattern, and printing a transparent mask sheet according to the chip mask pattern;

[0044] S2: male mold manufacturing; covering two layers of photosensitive dry film on the surface of the glass slide to form a glass slide with a photosensitive dry film layer, using a UV ultraviolet lamp to expose the glass slide using a transparent mask, thereby transferring the chip pattern on the mask to the photosensitive dry film layer by the principle of exposure imaging, then baking the exposed glass slide and developing it with a developing solution, and finally heating and curing to obtain a photosensitive dry film with a chip mask pattern as a male mold;

[0045] S3: microfluidic chip manufacturing: stirring the prepolymer and curing agent of PDMS to make the gas bubbles in the mixed solution uniform, vacuumizing the mixed solution through a vacuum dryer until the gas bubbles in the mixed solution disappear; then pouring the mixed solution into the male mold, placing it in an oven for curing; cutting the chip mask pattern to demold and punching holes at the inlet and outlet to form a chip embryo; finally, performing oxygen plasma treatment on the chip embryo, covering the photosensitive dry film side of the glass slide with a cover glass, bonding the two through a locking reaction to obtain a microfluidic chip.

[0046] Specifically, the ratio of prepolymer to curing agent is 10:1.

[0047] Specifically, the mixed solution is poured into the male mold, specifically, the edge of the male mold is wrapped with tin paper first, then placed in a culture dish, and then slowly poured into the prepared mixed solution to ensure that the PDMS is uniformly distributed in the male mold and the liquid level is level.

[0048] Specifically, the mixed solution is poured into the male mold and then vacuumized again.

[0049] Specifically, the oxygen plasma treatment on the chip embryo is performed by placing the cover glass and the glass slide in a plasma cleaner, with a radio frequency power of 45W and an oxygen plasma treatment time of 3min.

[0050] The dry film method is used to prepare the mold, which can manufacture channels with a size of less than 100 microns. The micro-machining technology that can be quickly carried out in a general laboratory can produce high-efficiency and low-cost microfluidic chips, which can be a useful supplement to traditional classic microfluidic processing technology and can provide an economical scientific research platform for the application of microfluidic chips.

[0051] The microfluidic chip based on droplet splitting and fusion and its manufacturing method according to the embodiment will be described in detail through specific examples.

[0052] The technical scheme adopted in the embodiment is as follows: a split-mixing type microfluidic concentration gradient droplet chip, comprising a glass bottom plate, wherein a PDMS substrate is arranged in the glass bottom plate, two continuous phase inlets and one dispersed phase inlet are arranged on one side of the top of the PDMS substrate, two continuous phase channels and a dispersed phase channel form a T-shaped structure to generate two groups of droplets. The output end of the T-shaped structure is provided with an inverted Y-shaped structure. The output end of the inverted Y-shaped structure is provided with a Y-shaped structure and a droplet fusion area.

[0053] With further limitation, the output end of the continuous phase inlet is arranged in a horizontal direction, and the output end of the dispersed phase inlet is arranged in a vertical direction.

[0054] With further limitation, the droplet fusion structure adopts an expansion channel structure.

[0055] With further limitation, the Y-shaped structure, the inverted Y-shaped structure and the droplet fusion structure are sequentially increased in expansion.

[0056] The beneficial effects of the embodiment are as follows: two T-shaped structures are arranged in the embodiment to simultaneously generate two groups of droplets, the droplets are split through the inverted Y-shaped structure design, and the droplet fusion is realized through the Y-shaped structure and the expansion structure. In the embodiment, the concentration gradient does not rely on diffusion mixing, but the diluent and the sample are divided into two groups of droplets A and B, the droplets are split into two groups when flowing through the bifurcation structure, and are A11, A22 and B11, B12. A11 meets B11 in the Y-shaped structure and is mixed in the expansion structure to generate a droplet string as a first concentration droplet. The first concentration droplet is split into two groups again when flowing through the bifurcation structure, one group meets A22 in the Y-shaped structure and is mixed to dilute into a second concentration droplet with a concentration smaller than the first concentration, and the other group meets B12 in the Y-shaped structure and is mixed to dilute into a third concentration droplet with a concentration greater than the first concentration. The chemical gradient distribution can control the step-by-step and convective distribution of the sample solution along the main flow. The key component microchannel of the step-by-step distribution is to the Y-shaped structure. By increasing the number of split-mixing structures, a complex and controllable gradient distribution can be generated.

[0057] Reference Figure 5 A preparation method of a split-mixing type microfluidic concentration gradient droplet chip, comprising the following steps:

[0058] S1: mask plate manufacturing; an AutoCAD program is used to create a mask, and a high-resolution transparent film mask sheet is printed by using a film printer;

[0059] S2: male mold making; after the glass slide is cleaned with anhydrous ethanol and deionized water and dried, two layers of photosensitive dry film are covered on the surface of the glass slide to prepare a dry film layer with a thickness of 60 μm, the glass slide with the photosensitive dry film layer is pressed at a temperature of 55°C and a pressure of 40 kpa for 1 min to complete the operation of removing bubbles and fixing the film, then the drawn microfluidic chip design is printed on a high-resolution transparent film as a mask, and then the glass slide is exposed to UV light, so that the microfluidic chip design pattern on the mask is transferred to the photosensitive dry film layer by the principle of exposure imaging, achieving the effect of fixing the channel shape, then the exposed glass slide is baked at a high temperature of 95°C for 4 min, then developed with a developing solution, and finally cured with a heating plate, and the remaining photosensitive dry film pattern is the male mold;

[0060] S3: microfluidic chip making: weighing PDMS prepolymer and curing agent: place a weighing paper on an electronic balance, then put a plastic cup into the balance, then zero the balance, pour the PDMS prepolymer into the plastic cup, and the pouring speed is from fast to slow until 18 g of PDMS prepolymer is obtained, then zero the balance, and then use a plastic dropper to drop the PDMS curing agent into the plastic cup until 1.8 g of PDMS curing agent is obtained, and the required ratio of prepolymer to curing agent is 10:1;

[0061] Stir until mixed evenly: use a rubber dropper or glass rod to stir the PDMS prepolymer and curing agent until the bubbles in the mixed solution are evenly distributed;

[0062] Vacuum: place the plastic cup in a vacuum dryer for vacuum operation, as bubbles will affect the observation of the situation in the microfluidic chip pipeline, so the bubbles in the mixed solution must be removed before the next step;

[0063] Pour the PDMS into the male mold: first wrap the edges of the male mold silicon wafer with a microchannel structure with tin paper, then place it in a culture dish to prevent PDMS from spilling during subsequent operations, then slowly pour the prepared PDMS into the culture dish to ensure that the PDMS on the male mold is evenly distributed and has good levelness, and the thickness is about 1 mm;

[0064] Vacuum: as bubbles may still be generated during pouring, a second vacuum operation is required, and the vacuum must be completely removed;

[0065] Baking: place the culture dish in an oven, set the temperature to 70°C, and the curing time is about 1.5 hours;

[0066] Cutting and demolding: use a scalpel to cut the microfluidic channel to the appropriate size, then demold, and use tweezers to clamp it into a culture dish;

[0067] Punching: Since the two-phase inlet and outlet of the microfluidic channel need to be punched to facilitate the insertion of the hose, the chip needs to be vertically punched using a PDMS manual puncher;

[0068] Bonding: The PDMS cover sheet and the glass slide with microfluidic channels are cleaned with anhydrous ethanol and deionized water and dried, and then the cover sheet and the glass slide are placed in a plasma cleaner, the selected radio frequency power is HIGH (about 45W), the oxygen plasma treatment time is 3min, after the oxygen plasma treatment, the glass slide and the PDMS cover sheet are taken out from the plasma cleaner, and then they are immediately pasted together by the oxygen plasma treated side using tweezers. Since the silicon hydroxyl groups on the surface of the substrate and the cover sheet can undergo condensation reaction when they come into contact, the condensation reaction can cause irreversible permanent bonding of the substrate and the cover sheet, and the microfluidic chip fabrication is completed.

[0069] In the S2, the stenciler is set to 55℃, 40kpa, and is pressed for 1min under this setting.

[0070] In the S3, the required ratio of the prepolymer and the curing agent is 10:1.

[0071] During the punching process, the puncher and the PDMS residue in the chip need to be completely cleaned out.

[0072] The technical effects of the embodiment are as follows:

[0073] 1. A microfabrication technology that can be quickly carried out in a general laboratory, thereby producing a high-efficiency, low-cost microfluidic chip, which can be a beneficial supplement to traditional classic microfluidic processing technology and can provide an economic research platform for the application of microfluidic chips, thereby promoting the popularization and rapid development of microfluidic technology to a certain extent.

[0074] 2. The embodiment is characterized in that it can generate small droplets using multi-phase microfluids and can regard each droplet as a micro-reactor. In addition, the droplet microfluidic technology can perform operations such as fusion, transportation, sorting, and splitting of droplets, and thus can be used in many research fields such as drug transportation, biochemical detection, cell culture, and specific virus screening.

[0075] 3. The preparation method mentioned in the embodiment is a beneficial supplement and attempt to traditional photolithography and precision machining processes, and can provide an economic research platform for more applications of microfluidic chips and promote the application of microfluidic chips in various fields.

[0076] 4. The embodiment can accurately control the liquid concentration in the spatial and temporal dimensions, thereby controlling the cell microenvironment, and has very promising application prospects in cell research.

[0077] 5、The embodiment has low requirement for the operator, high automation degree and stability, and increases controllability of the concentration gradient, which is beneficial to generate multi-component controllable concentration gradient.

[0078] The above merely describes the preferred embodiments of the present application, but the protection scope of the present application is not limited thereto, and any person skilled in the art can make equivalent replacements or changes to the technical scheme and the inventive concept of the present application within the technical scope disclosed by the present application, which should be covered within the protection scope of the present application.

Claims

1. A microfluidic chip based on droplet splitting and fusion, characterized by, The application relates to a microfluidic chip. The input part comprises a first input end (1), a second input end (2), a dispersed phase input end (3) and two T-shaped input structures, the longitudinal ends of the two T-shaped structures are connected with the dispersed phase input end (3), the first input end (1) and the second input end (2) are respectively connected with the first transverse ends of the two T-shaped input structures. The output part comprises a primary output structure and at least one secondary output structure, the primary output structure comprises two Y-shaped structures, the two Y-shaped structures are arranged in parallel and the main path ends of the two Y-shaped structures are respectively connected with the second transverse ends of the two T-shaped input structures, the adjacent branch path ends of the two Y-shaped structures form a primary mixing chamber (4) which is communicated with each other, and the primary mixing chamber (4) has a mixing output port. The secondary output structure comprises two end Y-shaped structures and at least one middle Y-shaped structure which is located between the two end Y-shaped structures, the end Y-shaped structures and the middle Y-shaped structure are arranged in parallel, the main path end of the end Y-shaped structure is communicated with the outer branch path end of the upper-stage Y-shaped structure, the main path end of the middle Y-shaped structure is communicated with the mixing output port of the primary mixing chamber (4), the branch path ends of the adjacent Y-shaped structures form secondary mixing chambers (5) which are communicated with each other, and the secondary mixing chambers (5) have mixing output ports. The mixing output port of the terminal secondary output structure and the outer branch path end of the end Y-shaped structure serve as a droplet output end (6). The two groups of droplets generated through the T-shaped structure are split into two groups of small droplets through the inverted Y-shaped structure and then are converged to the diffusion channel through the Y-shaped structure to be fused. The output part comprises a plurality of secondary output structures, the number of the middle Y-shaped structures of the plurality of secondary output structures gradually increases in the direction away from the primary output structure, the main path end of each end Y-shaped structure is communicated with the outer branch path end of the upper-stage end Y-shaped structure, and the main path end of the middle Y-shaped structure is communicated with the mixing output port of the upper-stage primary mixing chamber (4).

2. The droplet splitting and fusion based microfluidic chip according to claim 1, wherein, The Y-shaped structures of the output part are of the same size.

3. The droplet splitting and fusion based microfluidic chip according to claim 1, wherein, The application further discloses a microfluidic chip manufacturing method.

4. A method for fabricating a droplet splitting and fusion based microfluidic chip according to any one of claims 1-3, characterized in that, In S3, the ratio of the prepolymer and the curing agent is 10:

1. The application further discloses a microfluidic chip manufacturing method. In S3, the ratio of the prepolymer and the curing agent is 10:

1. ​ 5. The method of manufacturing according to claim 4, wherein, ​ 6. The method of claim 4, wherein, In S3, the mixed solution is poured into the male mold, specifically, the edge of the male mold is wrapped with tin paper first, then placed in a culture dish, and then slowly poured into the prepared mixed solution to ensure that the PDMS is evenly distributed in the male mold and the liquid level is level.

7. The method of manufacturing according to claim 6, wherein, In S3, the mixed solution is poured into the male mold and then vacuumized again.

8. The method of claim 4, wherein, In S3, the chip embryo is treated with oxygen plasma, specifically, the cover glass and the glass slide are placed in a plasma cleaner, the radio frequency power is 45W, and the oxygen plasma treatment time is 3min.

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