Handheld microfluidic sample processing device

Abstract

The utility model provides a handheld micro-fluidic sample processing device. The handheld micro-fluidic sample processing device comprises a clamp upper plate, a gas circuit connecting plate, a joint sealing plate, a chip sealing plate, a micro-fluidic chip and a clamp lower plate which are sequentially stacked from top to bottom, the upper clamp plate is provided with a hollowed-out area used for being connected with the second positioning hole of the lower clamp plate. The gas circuit connecting plate is provided with a plurality of gas circuit connecting ports which are densely arranged and first positioning holes which are connected with the clamp lower plate; the joint sealing plate is provided with a joint connecting port corresponding to the gas path connecting port of the gas path connecting plate and a third positioning hole for fixing the chip sealing plate; the chip sealing plate is provided with a liquid storage tank which is arranged corresponding to the joint connecting port of the joint sealing plate; the micro-fluidic chip comprises a chip main body and a reaction substrate, and a chip positioning area for placing the micro-fluidic chip is arranged on the clamp lower plate. According to the utility model, the hardware cost is reduced, and various functional operations can be satisfied.

Description

Technical Field

[0001] The utility model belongs to the technical field of microfluidics, and in particular relates to a handheld microfluidics sample processing device. Background Art

[0002] Microfluidics refers to the science and technology involved in systems that use microchannels (tens to hundreds of microns in size) to process or manipulate tiny fluids. It is an emerging interdisciplinary subject involving chemistry, fluid physics, microelectronics, new materials, biology, and biomedical engineering. It can integrate the process of analyzing samples in biology, chemistry, medicine, and other fields, including preparation, reaction, separation, detection, and other basic units, onto a micron-scale chip, and automatically complete the entire analysis process. Because of its miniaturization and integration, microfluidic devices are usually called microfluidic chips, also known as labs on a chip.

[0003] Microfluidic technology has the advantages of microquantity, high throughput, small sample requirements, and low reagent consumption, which makes it have very broad application prospects in different fields. In the field of in vitro diagnosis, it can play its own characteristics in IVD sub-fields such as biochemical analysis, immunodiagnosis, and molecular diagnosis, and has great potential to replace traditional IVD detection methods. The advantages in cell separation can greatly improve the detection rate and purity of circulating tumor cells, thereby providing key support for the treatment of related cancers. In the field of drug screening, such as stem cell chips, it can overcome the limitations of existing stem cells for in vitro research, and simulate the complex environment of stem cell growth and differentiation as much as possible by accurately controlling various factors in the stem cell microenvironment in real time. For example, a single-cell library preparation system (ZL202110977986.8) developed by Beijing Xunyin Biotechnology Co., Ltd. is used for cell labeling and library construction in single-cell sequencing, and the microfluidic chip and its droplet generation device (ZL201710364655.0) developed by Guangdong Yongnuo Medical Technology Co., Ltd. are used for the generation of droplets in digital PCR. The application of its microfluidic technology requires matching microfluidic chips and microfluidic control systems. The currently developed microfluidic operating systems have single functions and can only be used with a single chip for fixed projects. Different equipment is needed to carry out different projects. The hardware cost is high, the operation is complicated, and the convenience is poor. Utility Model Content

[0004] In order to solve the above problems, the utility model provides a handheld microfluidic sample processing device to achieve the purpose of matching with a variety of microfluidic chips, completing multiple projects with one device, reducing hardware costs, and meeting multiple functional operations.

[0005] In order to achieve the above purpose, the technical solution is as follows:

[0006] A handheld microfluidic sample processing device comprises a fixture upper plate, an air path connection plate, a joint sealing plate, a chip sealing plate, a microfluidic chip, and a fixture lower plate which are stacked in sequence from top to bottom;

[0007] The upper plate of the fixture is provided with a hollow area for passing the gas path connection port of the gas path connection plate, and is used to connect to the second positioning hole of the lower plate of the fixture;

[0008] The gas circuit connection plate is provided with a plurality of closely spaced gas circuit connection ports for connecting to an external gas source and a first positioning hole for connecting to the lower plate of the fixture;

[0009] The joint sealing plate is provided with a joint connection port corresponding to the gas path connection port of the gas path connection plate, and a third positioning hole for fixing the chip sealing plate;

[0010] The chip sealing plate is provided with a liquid storage tank corresponding to the joint connection port of the joint sealing plate, and a hook for clamping the third positioning hole of the joint sealing plate;

[0011] The microfluidic chip comprises a channel layer and a substrate layer, wherein the channel layer is tightly sealed with the chip sealing plate from top to bottom, and the substrate layer is tightly attached to the bottom of the channel layer;

[0012] The lower plate of the fixture is provided with a chip positioning area for placing the microfluidic chip, a first positioning pin for connecting the gas path connecting plate, and a second positioning pin for connecting the upper plate of the fixture. The lower plate of the fixture is connected to the upper plate of the fixture by a stainless steel buckle.

[0013] Furthermore, spring beads for forming point contact with the air path connecting plate are provided around the hollow area of ​​the upper plate of the clamp, and the second positioning holes are arranged at the four corners of the upper plate of the clamp; a buckle is provided on each side of the upper plate of the clamp; and the thickness of the upper plate of the clamp is less than or equal to 5 mm.

[0014] Furthermore, the gas path connecting plate also includes a connecting plate body, the connecting plate body is made of stainless steel or aluminum plate, and the thickness of the connecting plate body is 5-10mm; the gas path connecting ports are closely arranged in the middle position of the connecting plate body; the first positioning holes are arranged at the four corners of the connecting plate body.

[0015] Furthermore, the air circuit connection port is a pneumatic quick-connect connector, and the pneumatic quick-connect connector is threadedly connected to the air inlet on the connecting plate body.

[0016] Furthermore, the third positioning holes of the joint sealing plate are arranged at the four corners of the joint sealing plate; the joint sealing plate has one or more groups of joint connection ports, which are arranged corresponding to the liquid storage tanks of the chip sealing plate.

[0017] Furthermore, the liquid storage tank of the chip sealing plate is a cylinder, and the liquid storage tank is arranged in one or more groups; the liquid outlet of the liquid storage tank is a conical card hole, which forms a seal with the microfluidic chip; the top of the liquid storage tank is tightly attached to the joint sealing plate from top to bottom.

[0018] Furthermore, the channel layer of the microfluidic chip is located above the substrate layer and is tightly attached to the substrate layer; one or more groups of reaction units are arranged on the channel layer of the microfluidic chip; the reaction unit includes a channel and an opening, the opening of the reaction unit corresponds to the position of the liquid storage tank of the chip sealing plate up and down, and the channel of each reaction unit is connected to the corresponding opening; the size of the opening is 1-6mm.

[0019] Furthermore, the channel is a core reaction area, and its characteristic size is 10um-500um; the microfluidic chip material is one of polydimethylsiloxane or polycarbonate; the bonding method of the channel layer and the substrate layer is reversible bonding or irreversible bonding, and the bonding method is one of plasma bonding, chemical reagent bonding or biological tape bonding.

[0020] Furthermore, the lower plate of the clamp is a rectangular groove structure with an opening on one side, and a chip positioning area is set at the inner bottom of the lower plate of the clamp; the first positioning pins are set at the four corners of the inner bottom of the lower plate of the clamp, and the second positioning pins are set at the four corners of the top of the outer wall of the lower plate of the clamp; buckle grooves are provided on both sides of the lower plate of the clamp corresponding to the buckle positions of the upper plate of the clamp, and the thickness of the lower plate of the clamp does not exceed 5mm; an observation port is set in the middle of the lower plate of the clamp.

[0021] Compared with the prior art, the beneficial effects of the utility model are:

[0022] The device of the utility model has a small size, a simple structure, a low manufacturing cost, and is easy to operate. The modules can be assembled in sequence and it is easy to carry. Secondly, the device can be matched with microfluidic chips of different functional structures. The corresponding microfluidic chip can be selected according to the purpose. After assembly, an external gas source with a corresponding channel and pressure can be connected to realize the separation, mixing and packaging of different particles, solutions, and chemicals in the chip, providing a physical basis for subsequent biochemical reactions, and can be used for cell labeling in single-cell sequencing, droplet generation in digital PCR, preparation of gel microbeads, etc. In addition, the device of the utility model can adjust the gas access pressure and the amount of reaction liquid according to different projects to optimize the experimental conditions. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 It is a schematic diagram of the overall structure of the device of the utility model;

[0024] Figure 2 It is a structural exploded diagram of the device of the utility model;

[0025] Figure 3 This is a structural schematic diagram of the upper plate of the fixture of the utility model;

[0026] Figure 4 This is a structural schematic diagram of the gas path connecting plate of the utility model device;

[0027] Figure 5 This is a structural schematic diagram of a joint sealing plate of the utility model device;

[0028] Figure 6 This is a schematic diagram of the structure of the chip sealing plate of the device of the utility model;

[0029] Figure 7 It is a side view of the chip sealing plate of the device of the utility model;

[0030] Figure 8 This is a schematic diagram of the structure of chip A of the microfluidic chip of the utility model;

[0031] Fig. 9 This is a schematic diagram of the structure of the chip B of the microfluidic chip of the utility model;

[0032] Fig.10 This is a structural schematic diagram of the lower plate of the fixture of the utility model;

[0033] Fig.11 A microscope image of the PCR droplet generated in Example 2 of the present utility model;

[0034] Fig.12 This is a microscope image of the hydrogel microbeads of Example 3 of the utility model;

[0035] Fig.13 This is a graph showing the quality inspection results of the single-cell sequencing library of the control group of Example 4 of the present utility model;

[0036] Fig.14 This is a graph showing the quality inspection results of the single-cell sequencing library of the test group of Example 4 of the present utility model;

[0037] Fig.15 This is a graph showing the analysis results of single-cell sequencing data of the control group of Example 4 of the present utility model;

[0038] Fig.16 This is a graph of single-cell sequencing data analysis results for the test group of Example 4 of the present utility model.

[0039] Figure numerals: 1. upper plate of the fixture; 101. buckle; 102. spring top bead; 103. second positioning hole; 2. gas path connecting plate; 201. gas path connecting port; 202. first positioning hole; 3. joint sealing plate; 301. joint connecting port; 302. third positioning hole; 4. chip sealing plate; 401. liquid storage tank; 402. hook; 403. liquid outlet; 5. microfluidic chip; 501. channel layer; 502. opening; 503. substrate layer; 504. reaction unit; 505. channel; 6. lower plate of the fixture; 601. chip positioning area; 602. first positioning pin; 603. second positioning pin; 604. buckle groove; 605. observation port. DETAILED DESCRIPTION

[0040] The following will be combined with the drawings in the embodiments of the utility model to clearly and completely describe the technical solutions in the embodiments of the utility model. Obviously, the described embodiments are only part of the embodiments of the utility model, not all of the embodiments. Based on the embodiments in the utility model, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the utility model.

[0041] In the description of the present utility model, it should be noted that the orientation or position relationship indicated by the terms "center", "upper", "lower", "left", "right", "vertical", "horizontal", "inside", "outside", etc. is based on the orientation or position relationship shown in the drawings, which is only for the convenience of describing the present utility model and simplifying the description, and does not indicate or imply that the device or element referred to must have a specific orientation, be constructed and operated in a specific orientation, and therefore cannot be understood as a limitation of the present utility model; the terms "first", "second", and "third" are only used for descriptive purposes, and cannot be understood as indicating or implying relative importance. In addition, unless otherwise clearly specified and limited, the terms "installed", "connected", and "connected" should be understood in a broad sense, for example, it can be a fixed connection, a detachable connection, or an integral connection; it can be a mechanical connection or an electrical connection; it can be a direct connection, or it can be indirectly connected through an intermediate medium, or it can be the internal communication of two elements. For ordinary technicians in this field, the specific meanings of the above terms in the present utility model can be understood according to specific circumstances.

[0042] Example 1

[0043] A handheld microfluidic sample processing device, such as Figure 1-2 As shown, it includes a fixture upper plate 1, a gas path connection plate 2, a joint sealing plate 3, a chip sealing plate 4, a microfluidic chip 5, and a fixture lower plate 6 which are stacked in sequence from top to bottom;

[0044] like Figure 3As shown, the upper plate 1 of the fixture is provided with a hollow area for passing the gas path connection port 201 of the gas path connection plate 2, and a second positioning hole 103 for connecting the lower plate 6 of the fixture;

[0045] like Figure 4 As shown, the gas circuit connection plate 2 is provided with a plurality of closely spaced gas circuit connection ports 201 for connecting to an external gas source, and a first positioning hole 202 for connecting to the lower plate 6 of the fixture;

[0046] like Figure 5 As shown, the joint sealing plate 3 is provided with a closely spaced joint connection port 301 corresponding to the gas path connection port 201 of the gas path connection plate 2, and the joint connection port 301 connects the upper gas path connection plate 2 and the lower chip sealing plate 4; the joint sealing plate 3 is also provided with a third positioning hole 302 for fixing the chip sealing plate 4; in particular, the material of the joint sealing plate 3 includes but is not limited to silicone, fluororubber or rubber;

[0047] like Figure 6-7 As shown, the chip sealing plate 4 is provided with a liquid storage tank 401 corresponding to the joint connection port 301 of the joint sealing plate, and a hook 402 for clamping the positioning hole of the joint sealing plate;

[0048] The microfluidic chip 5 includes a channel layer 501 and a substrate layer 503. The channel layer 501 is tightly sealed with the chip sealing plate 4, and the substrate layer 503 is tightly attached to the bottom of the channel layer 501.

[0049] like Fig.10 As shown, the clamp lower plate 6 is provided with a chip positioning area 601 for placing the microfluidic chip 5, a first positioning pin 602 for connecting the gas path connecting plate 2, and a second positioning pin 603 for connecting the clamp upper plate 1. The clamp lower plate 6 is connected to the clamp upper plate 1 by a stainless steel buckle.

[0050] In this embodiment, spring beads for forming point contact with the air path connecting plate 2 are provided around the hollow area of ​​the upper plate 1 of the clamp, which are used to form point contact with the lower air path connecting plate 2 to avoid poor contact caused by processing flatness; second positioning holes 103 are arranged at the four corners of the upper plate 1 of the clamp; a buckle 101 is provided on each side of the upper plate 1 of the clamp, and the buckle 101 is preferably a stainless steel buckle 101, which is used to tightly buckle with the lower plate 6 of the clamp to form an overall seal of the device; the thickness of the upper plate 1 of the clamp does not exceed 5 mm.

[0051] In this embodiment, the gas circuit connecting plate 2 also includes a connecting plate body, which is made of stainless steel or aluminum plate, and has a thickness of 5-10 mm; the gas circuit connecting ports 201 are closely arranged in the middle of the connecting plate body; the first positioning holes 202 are arranged at the four corners of the connecting plate body;

[0052] In this embodiment, the air circuit connection port 201 is a pneumatic quick-connect connector, which is threadedly connected to the air inlet on the connecting plate body, and an external air source supplies the required pressure of the device through the pneumatic quick-connect connector.

[0053] In this embodiment, the liquid storage tank 401 of the chip sealing plate 4 is a cylinder, and the liquid storage tank 401 is arranged in one group or multiple groups. The storage volume of the liquid storage tank 401 is various and can store reaction liquids ranging from 10μL to 2000μL. The liquid outlet 403 at the bottom of the liquid storage tank 401 is provided with an arc to effectively avoid liquid residue. The liquid outlet 403 of the liquid storage tank 401 is a conical card hole, and the top of the liquid storage tank 401 is tightly attached to the joint sealing plate 3 up and down to ensure sealing.

[0054] In this embodiment, the channel layer 501 is located above the substrate layer 503 and is tightly attached to the substrate layer 503; the channel layer 501 is provided with one or more groups of reaction units 504; each reaction unit 504 includes a channel 505 and an opening 502, the opening 502 of the reaction unit corresponds to the position of the liquid storage tank of the chip sealing plate up and down, each reaction unit channel 505 is connected to the corresponding opening 502, the opening 502 corresponds to the position of the liquid storage tank 401 of the chip sealing plate 4 up and down, the size of the opening 502 is 1-6mm, which is convenient for connecting with the conical structure liquid outlet 403 of the liquid storage tank 401 of the chip sealing plate 4.

[0055] In this embodiment, the channel 505 is the core reaction area, and its characteristic size is 10um-500um, which is precisely processed by methods including but not limited to soft lithography; the microfluidic chip 5 is a material including but not limited to polydimethylsiloxane or polycarbonate; the bonding method of the channel layer 501 and the substrate layer 503 is reversible bonding or irreversible bonding, and the bonding method includes but is not limited to one of plasma bonding, chemical reagent bonding or biological tape bonding.

[0056] In particular, such as Figure 8-9 As shown, the microfluidic chip 5 is a chip A for digital PCR or gel beads preparation or a chip B for single-cell sequencing cell labeling, wherein chip A consists of 4 groups of reaction units 504, each group includes 3 openings 502, which are an oil phase pool, a reaction liquid pool, and a collection pool in sequence; chip B consists of 4 groups of reaction units 504, each group includes 4 openings 502, which are an oil phase pool, a cell suspension pool, a beads pool, and a collection pool in sequence.

[0057] In this embodiment, the lower plate 6 of the clamp is a rectangular groove structure with an opening on one side, and a chip positioning area 601 is set at the inner bottom of the lower plate 6 of the clamp; the first positioning pins 602 are set on the four corners of the inner bottom of the lower plate 6 of the clamp, and the second positioning pins 603 are set on the four corners of the top of the outer wall of the lower plate 6 of the clamp; buckle grooves 604 are provided on both sides of the lower plate 6 of the clamp corresponding to the buckle 101 positions of the upper plate 1 of the clamp, the thickness of the lower plate 6 of the clamp does not exceed 5 mm, and an observation port 605 is provided in the middle of the lower plate 6 to provide an observation window for the core area of ​​the microfluidic chip 5.

[0058] The present invention also discloses an assembly method of a handheld microfluidic sample processing device, which specifically includes the following steps:

[0059] Align the liquid outlet 403 of the liquid reservoir 401 of the chip sealing plate 4 with the opening 502 of the microfluidic chip 5 and seal tightly;

[0060] Align the joint connection port 301 of the joint sealing plate 3 with the liquid storage tank 401 of the chip sealing plate 4, and snap the hook 402 of the chip sealing plate 4 into the positioning hole of the joint sealing plate 3;

[0061] Place the microfluidic chip 5 on the chip positioning area 601 of the lower plate 6 of the fixture;

[0062] With the help of the first positioning pin 602 of the lower plate 6 of the fixture, gently insert the first positioning hole 202 of the gas circuit connection plate 2 into the first positioning pin 602 and fit it into the joint sealing plate;

[0063] The upper plate 1 of the fixture is fitted with the second positioning pin 603 of the lower plate 6 of the fixture through the second positioning hole 103 , and the microfluidic sample processing device is sealed by the buckle 101 .

[0064] The present invention also discloses a method for using a handheld microfluidic sample processing device, comprising the following steps:

[0065] Make or select a suitable microfluidic chip 5 according to the experimental purpose;

[0066] Assemble the microfluidic chip 5 and the chip sealing plate 4, add particles, solutions or chemicals into the liquid storage tank 401 of the chip sealing plate 4, and then assemble the microfluidic sample processing device as a whole;

[0067] Connect the external gas source through the gas connection plate 2, and set the gas source channel and the gas source pressure parameters of each channel according to the experimental purpose;

[0068] Run the microfluidic sample processing device and select an appropriate run time to obtain the target amount of mixture.

[0069] Example 2

[0070] This example is a specific example of the application of digital PCR detection. The specific operation steps are as follows:

[0071] (1) Reagents: Digital PCR premix, droplet generation oil, and analytical oil were purchased from Biorad, DNA samples extracted from PBMC were homemade, HER2 CNV primers and probes were purchased from Sangon Biotech (Shanghai) Co., Ltd., and DNase / RNase-Free Water was purchased from Thermo Fisher Scientific.

[0072] (2) Instruments and equipment: The droplet generation device used the handheld microfluidic sample processing device of the present invention, the digital PCR chip was a homemade PDMS chip, the external gas source had an accuracy of 1 mPa and a range of 20 mbar-2000 mbar, the PCR instrument (Mastercycler X50) was purchased from Eppendorf, and the droplet analyzer (QX200) and QuantaSoft software were purchased from Biorad;

[0073] (3) Prepare the digital PCR system: 2ul of PBMC DNA, 15ul of digital PCR premix, 1ul of HER2CNV primer, 0.8ul of probe, add nuclease-free water to 30ul, and mix well;

[0074] (4) Detection: Align the liquid outlet 403 of the liquid reservoir 401 of the chip sealing plate 4 with the opening 502 of the microfluidic chip 5 and fit them tightly together, add the digital PCR reaction system through the liquid reservoir 401, cover the joint sealing plate 3 for sealing, where the joint sealing plate 3 is preferably a silicone sealing plate, continue to assemble the gas path connecting plate 2, the clamp upper plate 1 and the clamp lower plate 6, and then fasten the device with the buckle 101, connect the external gas source, and operate at an air pressure of 250 mbar for the oil phase and 235 mbar for the water phase for 6 minutes to generate about 30,000 droplets;

[0075] (5) After the droplets are generated, Fig.11 As shown, the clamp was opened, and the droplets were aspirated from the collection wells into a 96-well plate for PCR, and PCR amplification was performed by denaturation at 95°C for 10 min, 95°C for 20 s, 60°C for 1 min, 40 cycles, 60°C for 5 min, and continuous storage at 4°C.

[0076] (6) Reading data: After the PCR is completed, take out the 96-well plate and assemble it in the fixture of the droplet reader. Then put it into the droplet reader, turn on the digital PCR reader, load the digital PCR droplet analysis oil, and perform detection and data analysis.

[0077] Example 3

[0078] This example is a specific example of the preparation of gel microbeads. The specific operation steps are as follows:

[0079] (1) Reagents: Acrylamide, methylenebisacrylamide, ammonium persulfate, Tris-HCl, EDTA, NaCl, and TEMED were purchased from Merck. Fluoride solution 7500 (Novec HFE-7500) was purchased from 3M. Surfactant (008-FluoroSurfactant-10G) was purchased from RNA Biotech. Acrydite-modified DNA primer design was purchased from Sangon Biotech (Shanghai) Co., Ltd.

[0080] (2) Instruments and equipment: The equipment adopts the handheld microfluidic sample processing device of the present invention, the microbead generation chip is a homemade PDMS chip, the main channel 505 is 60 μm wide, and the external gas source accuracy is 1 mPa;

[0081] The dispersed phase is composed of Tris-HCl pH 7.6, EDTA, NaCl, acrylamide, methylenebisacrylamide, ammonium persulfate and acrydite-modified DNA primers; the continuous phase is composed of fluorinated liquid 7500, TEMED and surfactants;

[0082] (3) Device assembly: Align the liquid outlet 403 of the liquid reservoir 401 of the chip sealing plate 4 with the opening 502 of the microfluidic chip 5 and fit them tightly together, add the dispersed phase and continuous phase mixed liquids into the liquid reservoir 401 respectively, cover with the joint sealing plate 3 to seal, continue to assemble the gas path connecting plate 2 and the clamp, fasten them, connect the external gas source, generate droplets into the centrifuge tube at a flow rate of 450 μL / h for the dispersed phase and 1000 μL / h for the continuous phase, and react at 65°C for 12 h to polymerize the gel beads;

[0083] (4) After the gel beads are generated, Fig.12 As shown, open the clamp and aspirate the gel beads from the collection well into a centrifuge tube for subsequent quality inspection and other applications.

[0084] The handheld microfluidic sample processing device of the present invention selects the corresponding microfluidic chip according to the purpose, and after assembly, an external gas source with a corresponding channel and pressure is connected to realize the separation, mixing and packaging of different particles, solutions and chemicals in the chip, providing a physical basis for subsequent biochemical reactions.

[0085] Example 4

[0086] This embodiment is applied in single-cell sequencing detection.

[0087] In this example, the single cell transcriptome detection reagent (Single cell RNA-Seq) of 10X Genomics was used to compare the performance of the Chromium single cell marker of 10X Genomics on the market, wherein the Chromium single cell marker served as the control group.

[0088] (1) Test reagents:

[0089]

[0090] (2) Experimental equipment:

[0091]

[0092] (3) Preparation of cell suspension and single cell labeling

[0093] Peripheral blood was drawn to obtain fresh PBMC cells, and the cell suspension was diluted to 1*10^6 cells / ml; the cells were labeled using the Chromium single cell labeling instrument and the device of the present invention, respectively, to obtain cell labeling products.

[0094] (4) Library preparation

[0095] The cell-labeled products were reverse transcribed, purified, and library prepared. The prepared library was detected for fragment distribution using the Agilent 2100 chip bioanalysis system.

[0096] (5) Sequencing and data analysis

[0097] The Illumina NovaSeq 6000 sequencer was used for sequencing, and the data such as the number of effective cells captured by the coded microbeads, the number of molecular tag UMIs, and the number of captured genes were analyzed. The results are as follows:

[0098]

[0099]

[0100] Conclusion: The test results are as follows Figure 13-16 It can be seen from the experimental analysis data that the microfluidic sample processing device of the present invention can achieve the same effect as the 10X Genomics product in the capture of cells, capture and analysis of mRNA molecules in single-cell transcriptome sequencing applications.

[0101] The utility model has a small size, simple structure, low manufacturing cost, simple operation, and can be assembled in order. It is easy to carry. Secondly, the device can be matched with microfluidic chips of different functional structures. The corresponding microfluidic chip can be selected according to the purpose. After assembly, the gas source of the corresponding channel and pressure can be connected to realize the separation, mixing and packaging of different particles, solutions and chemicals in the chip, providing a physical basis for subsequent biochemical reactions. In addition, the device of the present invention can adjust the gas access pressure and the amount of reaction liquid according to different projects to optimize the experimental conditions.

[0102] The above are only preferred embodiments of the present invention and do not limit the present invention in any way. Any technician in the relevant technical field, without departing from the scope of the technical solution of the present invention, may make any equivalent replacement or modification to the technical solution and technical content disclosed in the present invention, which shall be deemed as the content of the technical solution of the present invention and still fall within the protection scope of the present invention.

Claims

1. A handheld microfluidic sample processing device, characterized in that: It comprises a fixture upper plate (1), a gas path connection plate (2), a joint sealing plate (3), a chip sealing plate (4), a microfluidic chip (5), and a fixture lower plate (6) which are stacked in sequence from top to bottom; The clamp upper plate (1) is provided with a hollow area for passing the gas path connection port (201) of the gas path connection plate (2), and a second positioning hole (103) for connecting to the clamp lower plate (6); The gas path connection plate (2) is provided with a plurality of closely spaced gas path connection ports (201) for connection with an external gas source and a first positioning hole (202) for connection with the clamp lower plate (6); The joint sealing plate (3) is provided with a joint connection port (301) arranged corresponding to the gas path connection port (201) of the gas path connection plate (2), and a third positioning hole (302) for fixing the chip sealing plate (4); The chip sealing plate (4) is provided with a liquid storage tank (401) arranged corresponding to the joint connection port (301) of the joint sealing plate (3), and a hook (402) for clamping the third positioning hole (302) of the joint sealing plate (3); The microfluidic chip (5) comprises a channel (505) layer (501) and a substrate layer (503), wherein the channel (505) layer (501) is tightly sealed with the chip sealing plate (4) up and down, and the substrate layer (503) is tightly attached to the bottom of the channel (505) layer (501); The clamp lower plate (6) is provided with a chip positioning area (601) for placing the microfluidic chip (5), a first positioning pin (602) for connecting to the gas path connecting plate (2), and a second positioning pin (603) for connecting to the clamp upper plate (1). The clamp lower plate (6) is connected to the clamp upper plate (1) via a stainless steel buckle (101).

2. A handheld microfluidic sample processing device as claimed in claim 1, characterized in that: Spring beads (102) for forming point contact with the air path connecting plate (2) are arranged around the hollow area of ​​the clamp upper plate (1), and the second positioning holes (103) are arranged at the four corners of the clamp upper plate (1); a buckle (101) is arranged on each side of the clamp upper plate (1); and the thickness of the clamp upper plate (1) is less than or equal to 5 mm.

3. A handheld microfluidic sample processing device as claimed in claim 1, characterized in that: The gas path connecting plate (2) also includes a connecting plate body, the connecting plate body is made of stainless steel or aluminum plate, and the thickness of the connecting plate body is 5-10 mm; the gas path connecting ports (201) are closely arranged in the middle position of the connecting plate body; the first positioning holes (202) are arranged at the four corners of the connecting plate body.

4. A handheld microfluidic sample processing device as claimed in claim 3, characterized in that: The air circuit connection port (201) is a pneumatic quick-connect connector, and the pneumatic quick-connect connector is threadedly connected to the air inlet on the connecting plate body.

5. The handheld microfluidic sample processing device according to claim 1, characterized in that: The third positioning holes (302) of the joint sealing plate (3) are arranged at the four corners of the joint sealing plate (3); the joint sealing plate (3) has one or more groups of joint connection ports (301) which are arranged corresponding to the liquid storage tank (401) of the chip sealing plate (4).

6. A handheld microfluidic sample processing device as claimed in claim 1, characterized in that: The liquid storage tank (401) of the chip sealing plate (4) is a cylinder, and one or more groups of the liquid storage tanks (401) are provided. The liquid outlet (403) of the liquid storage tank (401) is a conical hole, which forms a seal with the microfluidic chip (5); the top of the liquid storage tank (401) is tightly attached to the joint sealing plate (3) up and down.

7. A handheld microfluidic sample processing device as claimed in claim 1, characterized in that: The channel (505) layer (501) of the microfluidic chip (5) is located above the substrate layer (503) and is tightly attached to the substrate layer (503); one or more reaction units (504) are arranged on the channel (505) layer (501) of the microfluidic chip (5); the reaction unit (504) comprises a channel (505) and an opening (502); the opening (502) of the reaction unit (504) corresponds to the position of the liquid storage tank (401) of the chip sealing plate (4) up and down; the channel (505) of each reaction unit (504) is connected to the corresponding opening (502); the size of the opening (502) is 1-6 mm.

8. A handheld microfluidic sample processing device as claimed in claim 7, characterized in that: The bonding method of the channel (505) layer (501) and the substrate layer (503) is reversible bonding or irreversible bonding, and the bonding method is one of plasma bonding, chemical reagent bonding or biological tape bonding.

9. The handheld microfluidic sample processing device according to claim 1, characterized in that: The lower plate (6) of the clamp is a rectangular groove structure with an opening on one side, and a chip positioning area (601) is set at the inner bottom of the lower plate (6); the first positioning pins (602) are set at the four corners of the inner bottom of the lower plate (6), and the second positioning pins (603) are set at the four corners of the top of the outer wall of the lower plate (6); buckle grooves (604) are provided on both sides of the lower plate (6) corresponding to the buckle (101) positions of the upper plate (1) of the clamp, and the thickness of the lower plate (6) does not exceed 5 mm; an observation port (605) is provided in the middle of the lower plate (6).

Images