Microfluidic Autoantibody Immunoassay Chip

Abstract

This application relates to a microfluidic autoantibody immunoassay chip, comprising: a top plate with a first through-hole for sample addition and a second through-hole for adding diluent; a middle plate with a sample addition area, a dilution area connected to the sample addition area, a mixing area, and a testing area connected to the mixing area, the testing area being able to detect the mixing effect; and a bottom plate including a labeling area connected to the testing area, a detection area connected to the labeling area, and a waste liquid area connected to the detection area. The inclusion of a testing area in this application to detect the mixing effect ensures the accuracy of the detection results. The ingenious connection of the mixing area, labeling area, and detection area on the chip, along with the synergistic effect of the control center, one-way valve, micropump, and motor, solves the problem of insufficient mixing of samples and diluents. The multiple sub-detection areas enable the detection of a large number of antigenic substances. This chip can be used to assist in the early screening and diagnosis of allergic diseases and autoimmune diseases.

Description

Technical Field

[0001] The present application belongs to the field of microfluidic immunoassay technology and relates to a microfluidic autoantibody immunoassay chip. Background Art

[0002] Microfluidic chips are one of the important components of chip laboratories. They are a technology that can accurately process trace amounts of liquids in channels with a width of micrometers. They have the advantages of reagent microquantity, high sensitivity, high throughput, and automation. Since the advent of microfluidic chips, they have been widely used in the medical field. Microfluidic chips only require a small amount of biological specimens or reagents. Their reaction system is small, the specific surface area is large, and the diffusion distance of reactants is short, which increases the chance of molecular interactions, shortens the reaction time, and has high sensitivity, and can measure low-concentration markers. Because it is easy to integrate and automate, it can eliminate cumbersome manual operations and reduce the chance of contamination. Its high-throughput nature allows multiple parallel tests to be performed on one chip at the same time, creating the possibility of auxiliary diagnosis of multiple clinical diseases. Moreover, this micro-analysis platform has a low manufacturing cost and a small size, and is easy to carry. Using this platform, medical workers can quickly complete the test in a resource-scarce medical environment without sending the specimen to a central laboratory.

[0003] Autoantibodies can be found in most autoimmune diseases, such as systemic lupus erythematosus and systemic sclerosis. In addition, there is evidence that autoantibodies are associated with cardiovascular diseases, chronic tumors and other diseases. Some autoantibodies are pathogenic, disease-specific and diagnostic. In the early or pre-disease stage of the disease, the concentration of autoantibodies will increase, so autoantibodies have early warning value; currently, many autoantibodies are used in the routine diagnosis and treatment of autoimmune diseases in clinical practice, which is of great value for the diagnosis, treatment monitoring and prognosis of autoimmune diseases. Due to technical limitations, most of the discovered autoantibodies are not used in routine clinical diagnosis. The development of new detection technologies with high specificity and sensitivity will help improve diagnostic performance and discover pathogenic autoantibodies that have not yet been identified. With the promotion of microfluidic immune chips, autoantibody detection has become one of the important research directions of microfluidic immune chips. Many problems still need to be overcome in the process of chip development, such as the problem of poor repeatability of test results due to uneven sample mixing. Summary of the invention

[0004] In order to solve the above-mentioned problems existing in the art, the present application aims to provide a microfluidic autoantibody immune detection chip, which tests the mixed solution after the sample and the diluent are mixed, and then performs subsequent reactions such as labeling and detection after the test confirms that they are fully mixed, to ensure the validity of the test results.

[0005] The technical solution of the present invention is as follows: a microfluidic autoantibody immune detection chip, comprising:

[0006] A top plate, wherein the top plate is provided with a first through hole for adding a sample and a second through hole for adding a diluent;

[0007] A middle plate, wherein the middle plate is provided with a sample adding area, a dilution area connected to the sample adding area, a mixing area, and a test area connected to the mixing area, wherein the test area can detect the mixing effect;

[0008] The bottom plate includes a marking area connected to the inspection area, a detection area connected to the marking area, and a waste liquid area connected to the detection area.

[0009] In some embodiments of the present application, the dilution zone and the mixing zone are connected via a first mixing pipe, a first micro pump is provided in the first mixing pipe, and the first mixing pipe has an irregular structure.

[0010] In some embodiments of the present application, a first stirring part and a second stirring part that do not contact each other are provided in the mixing zone.

[0011] In some embodiments of the present application, the mixing zone is connected to the inspection zone through a first flow channel, a first one-way valve and a second micropump are provided in the first flow channel, the other end of the inspection zone is connected to the second flow channel, the second flow channel is connected to a third flow channel and a fourth flow channel, the third flow channel is connected to another inlet of the mixing zone, a second one-way valve and a third micropump are provided in the third flow channel; the fourth flow channel is connected to the marking area located on the bottom plate, a third one-way valve and a fourth micropump are provided in the fourth flow channel.

[0012] In some embodiments of the present application, the outer shell of the inspection area is made of a transparent material, and the OD values ​​of multiple areas of the inspection area can be detected by a spectrophotometer.

[0013] In some embodiments of the present application, the labeling region is coated with a labeled antibody.

[0014] In some embodiments of the present application, the marking area is connected to the detection area via a fifth flow channel, and a fifth micro pump is disposed in the fifth flow channel.

[0015] In some embodiments of the present application, the detection area is provided with multiple sub-detection areas, the multiple sub-detection areas are coated with the same or different detection antibodies, the multiple sub-detection areas are provided with a detection card, the detection card has multiple detection strips, the multiple detection strips are coated with different detection antibodies, and the type and quantity of the detection antibodies and the labeled antibodies correspond one to one.

[0016] In some embodiments of the present application, the first one-way valve, the second one-way valve, the third one-way valve, the first micropump, the second micropump, the third micropump, the fourth micropump, the fifth micropump, the first motor, and the second motor are wirelessly connected to a control center.

[0017] In some embodiments of the present application, the top plate, the middle plate and the bottom plate are connected by snap-fitting.

[0018] The advantages of the present application are as follows: in the present application, the mixing effect is detected by setting up a test area, thereby ensuring the accuracy of the test result; the problem of insufficient mixing of the sample and the diluent in the test chip is solved by the clever connection of the mixing area, the labeling area, and the test area on the microfluidic chip and the coordinated action of the control center, the one-way valve, the micropump, and the motor; the setting of multiple sub-detection areas can detect a large number of antigen substances, and it is only necessary to coat the labeled antibody in the labeling area and the corresponding detection antibody in the detection area to detect a variety of antigen substances. The inclined setting of each microchannel (the first left microchannel, the first right microchannel...) ensures that the liquid can flow to the sub-detection areas away from the first sub-detection area under the action of gravity, and finally flow into the waste liquid area; the use of the microfluidic chip of the present application for immune detection can better solve the problem of poor repeatability of detection results caused by uneven sample mixing; the microfluidic autoantibody immune detection chip of the present application can not only be used to assist in the early screening and diagnosis of allergic diseases and autoimmune diseases, but also can be used for the detection of samples from various sources such as the environment and food, and the detectable molecular types are not limited to disease-related molecules. In fact, any antigen and antibody can be detected by this method, that is, its application is universal. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 This is a schematic diagram of the middle layer board of this application;

[0020] Figure 2 It is the schematic diagram of the top plate;

[0021] Figure 3 is a schematic diagram of the base plate;

[0022] Figure 4 for Figure 2 Enlarged view of point A in the middle;

[0023] Figure 5 for Figure 3 Cross-section at the middle BB;

[0024] Figure 6 is another schematic diagram of the first stirring blade;

[0025] Figure 7 is another schematic diagram of the first stirring blade;

[0026] Figure 8This is another schematic diagram of the first stirring blade.

[0027] In the figure: 1, top plate; 2, middle plate; 3, bottom plate; 4, first mixing pipe; 11, first through hole; 12, second through hole; 13, first observation window; 14, second observation window; 21, sample adding area; 22, dilution area; 23, mixing area; 24, inspection area; 31, marking area; 41, first vertical pipe; 42, first horizontal pipe; 43, second vertical pipe; 44, second horizontal pipe; 45, third vertical pipe; 46, third horizontal pipe 47, fourth vertical pipeline; 48, fourth horizontal pipeline; 49, fifth vertical pipeline; 410, fifth horizontal pipeline; 411, sixth vertical pipeline; 412, sixth horizontal pipeline; 413, seventh vertical pipeline; 51, first micro pump; 52, first one-way valve; 53, second micro pump; 54, second one-way valve; 55, third micro pump; 56, third one-way valve; 57, fourth micro pump; 58, fifth micro pump; 61, first rotating shaft; 62, first stirring blade; 63, first motor; 64, second rotating shaft; 65, second stirring blade; 66, second motor; 71, first flow channel; 72, second flow channel; 73, third flow channel; 74, fourth flow channel; 75, fifth flow channel; 81, first sub-detection area; 82, first left micro-channel; 83, first right micro-channel; 84, second left sub-detection area; 85, second right sub-detection area; 86, second left micro-channel; 87, third left micro-channel; 88, first Four left micro-channels; 89, third left sub-detection area; 810, fourth left sub-detection area; 811, fifth left sub-detection area; 812, second right micro-channel; 813, third right micro-channel; 814, fourth right micro-channel; 815, third right sub-detection area; 816, fourth right sub-detection area; 817, fifth right sub-detection area; 818, detection card; 819, fifth left micro-channel; 820, fifth right micro-channel; 91, left waste liquid area; 92, right waste liquid area. DETAILED DESCRIPTION

[0028] The following examples are provided to illustrate the present invention in more detail, but they do not constitute a limitation to the present invention. The experimental methods in the following examples are conventional methods unless otherwise specified. The reagents used in the following examples are commercially available common reagents unless otherwise specified.

[0029] See also Figure 1 An embodiment of the present application provides a microfluidic autoantibody immune detection chip, comprising: a top plate 1, on which a first through hole 11 for adding a sample and a second through hole 12 for adding a diluent are provided; a middle plate 2, on which a sample adding area 21, a dilution area 22 connected to the sample adding area 21, a mixing area 23, and a test area 24 connected to the mixing area 23 are provided; a bottom plate, which includes a marking area 31 connected to the test area 24, a detection area interconnected with the marking area 31, and a waste liquid area connected to the detection area.

[0030] The top plate 1 is provided with a first observation window 13 and a second observation window 14 . The first observation window 13 covers the sample adding area 21 and the dilution area 22 , through which the sample adding area 21 and the dilution area 22 can be viewed. The second observation window 14 covers the mixing area 23 , through which the mixing area 23 can be viewed.

[0031] The first through hole 11 is connected to the sample adding area 21 , and the second through hole 12 is connected to the dilution area 22 .

[0032] In order to prevent the sample from contacting the external environment, the first through hole 11 may be provided with an open or closed cover. After the cover is opened, the sample is added to the sample adding area 21 through the first through hole 11, and then the cover is closed, so that the sample and the external environment are isolated from each other; similarly, the second through hole 12 may also be provided with an open or closed cover to prevent the sample from contacting the external environment.

[0033] In this embodiment, the first through hole 11 is closer to the second observation window 14 than the second through hole 12 .

[0034] Similarly, on the middle plate 2, the sample adding area 21 is closer to the mixing area 23 than the dilution area 22. In order to improve the mixing effect, the dilution area 22 and the mixing area 23 are connected through the first mixing pipe 4. The first micro pump 51 is provided in the first mixing pipe 4. The first mixing pipe 4 is an irregular S-shaped structure.

[0035] Further, the first mixing pipe 4 includes a plurality of interconnected transverse pipes and vertical pipes. Specifically, one end of the first vertical pipe 41 is connected to the sample adding area 21, the other end of the first vertical pipe 41 is connected to the first transverse pipe 42, the other end of the first transverse pipe 42 is connected to one end of the second vertical pipe 43, the second vertical pipe 43 extends toward the mixing area 23, the other end of the second vertical pipe 43 is connected to the second transverse pipe 44, the other end of the second transverse pipe 44 is connected to the third vertical pipe 45, the third vertical pipe 45 extends in the opposite direction to the second vertical pipe 43, the third vertical pipe 45 extends toward the sample adding area 21, the other end of the third vertical pipe 45 is connected to the third transverse pipe 45, and the third vertical pipe 45 extends in the opposite direction to the second vertical pipe 43. The first transverse pipe 42 and the third transverse pipe 46 are connected to each other at the same time, and the other end of the third transverse pipe 46 is connected to the fourth vertical pipe 47, the other end of the fourth vertical pipe 47 is connected to the fourth transverse pipe 48, the other end of the fourth transverse pipe 48 is connected to the fifth vertical pipe 49, the fifth vertical pipe 49 extends to the mixing zone 23, the other end of the fifth vertical pipe 49 is connected to the fifth transverse pipe 410, the other end of the fifth transverse pipe 410 is connected to the sixth vertical pipe 411, the sixth vertical pipe 411 extends to the mixing zone 23, the other end of the sixth vertical pipe 411 is connected to the sixth transverse pipe 412, the sixth transverse pipe 412 is connected to the seventh vertical pipe 413, and the seventh vertical pipe 413 is connected to the mixing zone 23. The pipe walls of the first transverse pipe 42 and the third transverse pipe 46 are wavy, which can play a better blocking effect and improve the mixing efficiency compared with the straight pipe.

[0036] The other end of the first mixing pipe 4 is connected to the mixing zone 23, and a first stirring part and a second stirring part which are not in contact with each other are provided in the mixing zone 23. The first stirring part includes a first rotating shaft 61, a first stirring blade 62, and a first motor 63. The first rotating shaft 61 is located at the center of the first stirring part. The stirring blade is connected to the first rotating shaft 61. The stirring blade is symmetrically arranged about the first rotating shaft 61. The first motor 63 is connected to the first rotating shaft 61 to drive the first rotating shaft 61 to rotate; the second stirring part includes a second rotating shaft 64, a second stirring blade 65, and a second motor 66. The second rotating shaft 64 is located at the center of the second stirring part. The stirring blade is connected to the second rotating shaft 64. The stirring blade is symmetrically arranged about the second rotating shaft 64; the second motor 66 is connected to the second rotating shaft 64 to drive the second rotating shaft 64 to rotate.

[0037] Furthermore, in order to improve the stirring efficiency of the stirring blade, the first stirring blade 62 may be a spindle-shaped structure with a thick middle and thin ends (see Figure 6), so that when the stirring blade is working, the middle thick part drives the liquid to move with a greater amplitude than the two thin parts, so that the liquid close to the middle of the stirring blade will flow to both sides, and the liquid away from the stirring blade will flow to the middle of the stirring blade, so that the mixing effect can be further improved; the first stirring blade 62 can also be a structure with one side straight and the other side wavy, and one side of the wavy line structure faces the direction of rotation of the stirring blade (see Figure 7 ); The first stirring blade 62 may also be a thin structure in the middle and thick at both ends (see Figure 8 ).

[0038] The mixing area 23 is connected with the inspection area 24 through the first flow channel 71, and the first flow channel 71 is provided with a first one-way valve 52 and a second micro pump 53. The other end of the inspection area 24 is connected with the second flow channel 72, and the second flow channel 72 is connected with the third flow channel 73 and the fourth flow channel 74, wherein the third flow channel 73 is connected with another inlet of the mixing area 23, and the third flow channel 73 is provided with a second one-way valve 54 and a third micro pump 55; wherein the fourth flow channel 74 is connected with the marking area 31 located on the bottom plate, and the fourth flow channel 74 is provided with a third one-way valve 56 and a fourth micro pump 57.

[0039] In order to be able to test the mixing effect, the present embodiment is provided with a test area 24. The outer shell of the test area 24 is made of transparent material. The OD values ​​of multiple areas of the test area 24 can be tested by a spectrophotometer. If there is a large difference, it means that the mixing is uneven. It is necessary to open the second one-way valve 54 and the third micro pump 55, and pump the mixed liquid in the test area 24 back into the mixing area 23, stir and mix, and then test it by a spectrophotometer. In this way, the mixing effect can be guaranteed.

[0040] The labeling region 31 is coated with a labeled antibody, and further, the labeled antibody is an autoantibody target detection object. Optionally, the autoantibody target detection object is selected from dsDNA, AnuA, anti-Ku, AHA, ssDNA, SM, RNP, RIB, AMA-M2, Jo-1, SSA or SSB. It can be understood that the coated labeled antibody is not limited thereto and can be adjusted according to the detection target substance.

[0041] Specifically, the labeled antibody in the labeling area 31 combines with the antigen in the mixed solution to form a binary complex. The labeling area 31 is connected to the detection area through the fifth flow channel 75, and the fifth micro pump 58 is disposed in the fifth flow channel 75.

[0042] The detection area is provided with a plurality of sub-detection areas, and the plurality of sub-detection areas are coated with the same or different detection antibodies. The sub-detection area directly connected to the fifth flow channel 75 is the first sub-detection area 81. The first left micro-channel 82 and the first right micro-channel 83 are provided on both sides of the first sub-detection area 81. The first left micro-channel 82 and the first right micro-channel 83 are arranged obliquely. The connection between the first left micro-channel 82 and the first right micro-channel 83 and the first sub-detection area 81 is higher than any other part of the first left micro-channel 82 and the first right micro-channel 83. The first left micro-channel 82 is connected to the second left sub-detection area 84, and the first right micro-channel 83 is connected to the second right sub-detection area 85. That is, the second left sub-detection area 84 and the second right sub-detection area 85 are connected. The height of the zone 85 is lower than that of the first sub-detection zone 81; a second left micro-channel 86 is provided on the other side of the second left sub-detection zone 84 opposite to the first left micro-channel 82, the second left micro-channel 86 is inclined, and the connection between the second left micro-channel 86 and the second left sub-detection zone 84 is higher than any other part of the second left micro-channel, and the second left micro-channel 86 is connected to the third left sub-detection zone 89, that is, the height of the third left sub-detection zone 89 is lower than that of the second left sub-detection zone 84; a third left micro-channel 87 is provided on the other side of the third left sub-detection zone 89 opposite to the second left micro-channel 86, the third left micro-channel 87 is inclined, and the connection between the third left micro-channel 87 and the third left sub-detection zone 89 is higher than any other part of the third left micro-channel, and the third left micro-channel 87 is inclined. The third left micro-channel 87 is connected to the fourth left sub-detection area 810, that is, the height of the fourth left sub-detection area 810 is lower than the third left sub-detection area 89; the fourth left micro-channel 88 is provided on the other side of the fourth left sub-detection area 810 opposite to the third left micro-channel 87, and the fourth left micro-channel 88 is inclined. The connection between the fourth left micro-channel 88 and the fourth left sub-detection area 810 is higher than any other part of the fourth left micro-channel, and the fourth left micro-channel 88 is connected to the fifth left sub-detection area 811, that is, the height of the fifth left sub-detection area 811 is lower than the fourth left sub-detection area 810; similarly, the second right micro-channel 812 is provided on the other side of the second right sub-detection area 85 opposite to the first right micro-channel 83, and the second right micro-channel 81 2 is arranged obliquely, the connection between the second right micro-channel 812 and the second right sub-detection area 85 is higher than any other part of the second right micro-channel, and the second right micro-channel 812 is connected to the third right sub-detection area 815, that is, the height of the third right sub-detection area 815 is lower than the second right sub-detection area 85; the other side of the third right sub-detection area 815 opposite to the second right micro-channel 812 is provided with a third right micro-channel 813, the third right micro-channel 813 is arranged obliquely, the connection between the third right micro-channel 813 and the third right sub-detection area 815 is higher than any other part of the third right micro-channel, and the third right micro-channel 813 is connected to the fourth right sub-detection area 816, that is, the height of the fourth right sub-detection area 816 is lower than the third right sub-detection area 815;A fourth right micro-channel 814 is provided on the other side of the fourth right sub-detection area 816 opposite to the third right micro-channel 813. The fourth right micro-channel 814 is inclined. The connection between the fourth right micro-channel 814 and the fourth right sub-detection area 816 is higher than any other part of the fourth right micro-channel 814. The fourth right micro-channel 814 is connected to the fifth right sub-detection area 817, that is, the height of the fifth right sub-detection area 817 is lower than that of the fourth right sub-detection area 816.

[0043] A detection card 818 is provided in the plurality of sub-detection areas, and a plurality of detection strips are provided in the detection card 818. The plurality of detection strips are coated with different detection antibodies, and the types and quantities of the detection antibodies and the labeled antibodies correspond one to one. The binary complex object in the detection area forms a ternary complex with the detection antibody on the detection strip.

[0044] To facilitate waste liquid collection, the detection area also includes a left waste liquid area 91 and a right waste liquid area 92. The left waste liquid area 91 is connected to the fifth left sub-detection area 811 through the fifth left micro-channel. Similarly, the fifth left micro-channel 819 is inclined, and the connection between the fifth left micro-channel and the fifth left sub-detection area 811 is higher than any other part of the fifth left micro-channel 819, that is, the height of the left waste liquid area 91 is lower than the fifth left sub-detection area 811; correspondingly, the right waste liquid area 92 is connected to the fifth right sub-detection area 817 through the fifth right micro-channel. Similarly, the fifth right micro-channel 820 is inclined, and the connection between the fifth right micro-channel 820 and the fifth right sub-detection area 817 is higher than any other part of the fifth right micro-channel, that is, the height of the right waste liquid area 92 is lower than the fifth right sub-detection area 817.

[0045] Wherein the detection antibody is an autoantibody target detection object. Optionally, the detection antibody target detection object is selected from dsDNA, AnuA, anti-Ku, AHA, ssDNA, SM, RNP, RIB, AMA-M2, Jo-1, SSA or SSB. It can be understood that the coated detection antibody is not limited thereto and can be adjusted according to the detection target substance.

[0046] In the present application, the one-way valve, micro pump, motor and control center are connected wirelessly, and the one-way valve, micro pump and motor work under the drive of the control center.

[0047] In the present application, the top plate 1, the middle plate 2 and the bottom plate are connected by snap-fitting, wherein the inspection area 24 on the middle plate 2 is separately arranged and does not overlap with the areas on the top plate 1 and the bottom plate; in addition to the above-mentioned main structures, the top plate 1, the middle plate 2 and the bottom plate of the detection chip of the present application also have many air holes and make-shift holes and columns for assembly and fixation.

[0048] This embodiment also provides a method for using the above detection chip, as follows:

[0049] The sample is added through the first through hole, and the diluent is added through the second through hole. The diluent drives the sample to flow into the first mixing pipe, and the first micro pump is turned on. Under the drive of the first micro pump, the mixed liquid consisting of the sample and the diluent flows into the mixing area. Under the drive of the first motor and the second motor, the first stirring part and the second stirring part stir the mixed liquid. After the stirring is completed, the first one-way valve and the second micro pump are opened, and the mixed liquid enters the inspection area. The mixed liquid in the inspection area is tested at multiple points by a spectrophotometer. If the OD value difference is large, the second one-way valve and the third micro pump are opened, and the mixed liquid re-enters the mixing area for mixing; after mixing, it enters the inspection area to detect the OD value; if the OD values ​​are the same or approximately the same, the third one-way valve and the third micro pump are opened. The mixed liquid enters the marking area along the fourth flow channel, forms an antigen-antibody binary complex with the labeled antibody in the marking area, and the binary complex enters the detection area along the fifth flow channel, passes through the first sub-detection area, and then flows into the second left detection area, the second right detection area...the fifth right detection area arranged in the first sub-detection area in sequence. The content of the analyte in the sample is calculated by detecting the amount of the ternary complex in the detection area, and the mixed liquid that is not combined with the detection antibody flows into the left waste liquid area and the right waste liquid area. The quality control marker specifically combines with the quality control antibody on the quality control line to form a quality control marker-quality control antibody complex. The effectiveness of the detection chip is determined by detecting the quality control marker-quality control antibody complex in the detection area.

[0050] In the present application, the mixing effect is detected by setting up a test area to ensure the accuracy of the test results; the problem of insufficient mixing of samples and diluents in the test chip is solved by the clever connection of the mixing area, labeling area, and detection area on the microfluidic chip and the coordinated action of the control center, one-way valve, micropump, and motor; the setting of multiple sub-detection areas can detect a large number of antigen substances. It only needs to coat the labeled antibody in the labeling area and the corresponding detection antibody in the detection area to detect a variety of antigen substances. The inclined setting of each microchannel (the first left microchannel, the first right microchannel...) ensures that the liquid can flow to the sub-detection areas away from the first sub-detection area under the action of gravity, and finally flow into the waste liquid area; the use of the microfluidic chip of the present application for immune detection can better solve the problem of poor repeatability of detection results caused by uneven sample mixing; the microfluidic autoantibody immune detection chip of the present application can not only be used to assist in the early screening and diagnosis of allergic diseases and autoimmune diseases, but also can be used for the detection of samples from various sources such as the environment and food, and the detectable molecular types are not limited to disease-related molecules. In fact, any antigen and antibody can be detected by this method, that is, its application is universal.

[0051] When the terms "vertical", "horizontal", "left", "right", "up", "down", "inside", "outside", "bottom" and the like are used to indicate an orientation or positional relationship, it is based on the orientation or positional relationship shown in the drawings and is only for the convenience of description, and does not indicate or imply that the device or element referred to must have a specific orientation, be constructed and operate in a specific orientation, and therefore cannot be understood as a limitation of the present application. In addition, the terms "first", "second", etc. are used for descriptive purposes only and cannot be understood as indicating or implying relative importance. The term "and / or" includes any and all combinations of one or more related listed items.

[0052] The technical features of the above-described embodiments may be arbitrarily combined. To make the description concise, not all possible combinations of the technical features in the above-described embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0053] The above-described embodiments only express several implementation methods of the present application, and the descriptions thereof are relatively specific and detailed, but they cannot be construed as limiting the scope of the patent. It should be noted that, for a person of ordinary skill in the art, several modifications and improvements can be made without departing from the concept of the present application, and these all belong to the protection scope of the present application. Therefore, the protection scope of the present application shall be subject to the attached claims.

Claims

1. A microfluidic autoantibody immune detection chip, characterized in that: include: A top plate, wherein the top plate is provided with a first through hole for adding a sample and a second through hole for adding a diluent; A middle plate, wherein the middle plate is provided with a sample adding area, a dilution area connected to the sample adding area, a mixing area, and a test area connected to the mixing area, wherein the test area can detect the mixing effect; A bottom plate, the bottom plate comprising a marking area connected to the inspection area, a detection area connected to the marking area, and a waste liquid area connected to the detection area; The dilution zone is connected to the mixing zone through a first mixing pipe, a first micro pump is arranged in the first mixing pipe, the first mixing pipe is an irregular structure, and the first mixing pipe includes a plurality of interconnected transverse pipes and vertical pipes; The mixing area is connected with the inspection area through a first flow channel, a first one-way valve and a second micro pump are arranged in the first flow channel, the other end of the inspection area is connected with the second flow channel, the second flow channel is connected with the third flow channel and the fourth flow channel, the third flow channel is connected with another entrance of the mixing area, a second one-way valve and a third micro pump are arranged in the third flow channel; the fourth flow channel is connected with the marking area located on the bottom plate, a third one-way valve and a fourth micro pump are arranged in the fourth flow channel; The shell of the inspection area is made of transparent material, and the OD values ​​of multiple areas of the inspection area can be detected by a spectrophotometer; The marking area is connected to the detection area through a fifth flow channel, and a fifth micro pump is provided in the fifth flow channel; The detection area is provided with multiple sub-detection areas, the multiple sub-detection areas are connected by multiple micro-channels, the multiple micro-channels are arranged obliquely, the multiple sub-detection areas are coated with the same or different detection antibodies, the multiple sub-detection areas are provided with detection cards, the detection card has multiple detection strips, the multiple detection strips are coated with different detection antibodies, and the types and quantities of the detection antibodies and the labeled antibodies correspond one to one.

2. The microfluidic autoantibody immune detection chip according to claim 1, characterized in that: The mixing zone is provided with a first stirring portion and a second stirring portion which are not in contact with each other.

3. The microfluidic autoantibody immune detection chip according to claim 1, characterized in that: The labeling area is coated with a labeling antibody.

4. The microfluidic autoantibody immune detection chip according to claim 1, characterized in that: The first one-way valve, the second one-way valve, the third one-way valve, the first micropump, the second micropump, the third micropump, the fourth micropump, the fifth micropump, the first motor, the second motor and the control center are wirelessly connected.

5. The microfluidic autoantibody immune detection chip according to claim 1, characterized in that: The top plate, the middle plate and the bottom plate are connected by snapping.

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