Microfluidic single-molecule immunoassay analyzer

Abstract

The utility model discloses a micro-fluidic single molecule immunity analyzer, which comprises a micro-fluidic chip, a motion control module, a temperature control module, a pressing module, a magnetic control module and a signal acquisition module, and the micro-fluidic chip comprises at least one group of sample area, reagent area, acquisition area and waste liquid area; the sample area and the reagent area are communicated to the acquisition area through the test micro-channel, the acquisition area is communicated with the waste liquid area, the sample area is used for adding a certain amount of to-be-tested samples, the reagent area contains magnetic beads coated with antibodies, and the motion control module is used for driving the micro-fluidic chip to sequentially pass through the pressing module, the magnetic control module and the signal acquisition module. The pressing module is used for adding a first diluent into the sample area and adding a second diluent into the reagent area, and the magnetic control module is used for adsorbing an antibody on a magnetic bead into the collection area by combining with an antigen to be detected. The device has the advantages of few used samples, high detection speed, simplicity in operation and compact structure.

Description

Technical Field

[0001] The utility model relates to the technical field of photovoltaics, in particular to a microfluidic single molecule immunoassay analyzer. Background Art

[0002] Common immunoassay methods currently include radioimmunoassay, enzyme-linked immunosorbent assay, colloidal gold immunoassay, fluorescence immunoassay, chemiluminescence immunoassay, and PCR immunoassay. However, detection instruments designed based on these methodologies often suffer from issues such as insufficient sensitivity, low detection limits, large overall size, high cost, and long reaction times. For example, detection instruments designed or modified based on traditional IVD (in vitro diagnostic technology) or POCT (point-of-care) instruments often include basic components such as cuvette handling, a robotic arm, reagent probes, sample probes, a fluidics system, sample loading and disposal modules, reagent cryogenic storage, and incubation reaction systems. This results in a complex overall structure, large size, and high cost. Utility Model Content

[0003] Based on this, it is necessary to provide a microfluidic single molecule immunoassay analyzer. The microfluidic single molecule immunoassay analyzer of the present invention has one or more advantages of a more compact structure, light weight, lower overall cost, higher detection sensitivity, and faster reaction speed.

[0004] An embodiment of the present application provides a microfluidic single-molecule immunoassay analyzer.

[0005] A microfluidic single-molecule immunoassay analyzer comprises a microfluidic chip, a motion control module, a temperature control module, a pressing module, a magnetic control module and a signal acquisition module, wherein the microfluidic chip comprises a sample area, a reagent area, a collection area and a waste liquid area, the sample area and the reagent area are connected to the collection area through a test microchannel, the collection area is connected to the waste liquid area, the sample area is used to add a certain amount of sample to be tested, the reagent area contains magnetic beads coated with antibodies, the collection area is connected to antibodies and can achieve adsorption of magnetic beads, the waste liquid area is used to collect free magnetic beads that are not adsorbed, and the motion control module is used to drive the microfluidic chip. The fluidic chip passes through the pressing module, the magnetic control module and the signal acquisition module in sequence. The pressing module is used to add the first diluent into the sample area to realize a mixed incubation reaction of the sample to be tested, the whole blood treatment reagent and the first diluent in the sample area to form the antigen to be tested. The pressing module is also used to add the second diluent into the reagent area to realize the dissolution of the freeze-dried antibody-coated magnetic beads in the reagent area into a liquid state. The magnetic control module is used to realize that the antibodies on the magnetic beads are adsorbed in the acquisition area by binding to the antigen to be tested. The acquisition module is used to detect the number of magnetic beads in the acquisition area. The temperature control module is used to control the test temperature.

[0006] In some embodiments, the sample area is pre-loaded with whole blood processing reagents.

[0007] In some embodiments, the microfluidic chip further includes a first diluent area, which is connected to the sample area to enable the addition of a first diluent into the sample area, thereby enabling the test sample, whole blood processing reagent, and the first diluent in the sample area to be mixed and incubated to form the test antigen.

[0008] In some embodiments, the microfluidic chip further includes a second diluent area, which is connected to the reagent area to enable the addition of a second diluent into the reagent area, thereby dissolving the freeze-dried antibody-coated magnetic beads in the reagent area into a liquid state.

[0009] In some embodiments, the microfluidic chip includes at least one test group and at least one reference group, and the test group and the reference group both include the sample area, the reagent area, the collection area and the waste liquid area. The sample area and the reagent area in the reference group are connected to the corresponding collection area through a reference microchannel, and the reference microchannel has the same length as the test microchannel. The sample area in the reference group contains an artificially synthesized antigen to be tested, and the first diluent and the second diluent in the test group and the reference group are added synchronously through the same pressing module.

[0010] In some embodiments, the pressing module includes a driving component and at least two pressing heads, and the driving component is connected to the pressing head to drive the pressing head to press synchronously. When the motion control module drives the microfluidic chip to move to the first diluent area located at the pressing module, the driving component drives the pressing head to press synchronously to add the first diluent in the first diluent area to the sample area; when the motion control module drives the microfluidic chip to move to the second diluent area located at the pressing module, the driving component drives the pressing head to press synchronously to add the second diluent in the second diluent area to the reagent area.

[0011] In some embodiments, the magnetic control module includes a first electromagnet and a second electromagnet, the first electromagnet and the second electromagnet are spaced apart in the vertical direction, the motion control module is capable of driving the microfluidic chip to move between the first electromagnet and the second electromagnet, and the magnetic poles of the first electromagnet and the second electromagnet are capable of alternating;

[0012] When the first electromagnet located at the bottom is working, it attracts the magnetic beads and accelerates them to move downward. The magnetic beads bound to the antigen bind to the antibodies coated on the surface of the chip, and the magnetic beads not bound to the antigen are magnetically sucked downward; when the second electromagnet located at the top is working, the magnetic force of the second electromagnet is controlled to be greater than the gravity of the magnetic beads, and the magnetic beads not fixed on the chip are sucked upward and separated from the bottom of the chip and enter the collection area.

[0013] In some embodiments, the magnetic control module further includes an air pressure pulse component, which is used to apply air pressure pulses to the collection area. When the first electromagnet located below is working, the collection area is driven by the air pressure pulse to move the magnetic beads back and forth, so that they are evenly distributed in the collection area and reduce aggregation or stacking. When the second electromagnet located above is working, the collection area is driven by the air pressure pulse to accelerate the magnetic beads out of the collection area.

[0014] And / or, the magnetic pole area of ​​the first electromagnet is 2 to 5 times larger than the area of ​​the collection area;

[0015] And / or, the magnetic pole area of ​​the second electromagnet is 2 to 5 times larger than the area of ​​the collection area.

[0016] In some embodiments, the signal acquisition module includes an LED white light source, a light doffer, and an image sensor;

[0017] The light homogenizing plate and the image sensor are spaced apart in the vertical direction, and the microfluidic chip can pass between the light homogenizing plate and the image sensor; the LED white light source is used to provide bright field imaging illumination, the light homogenizing plate is used to homogenize the LED white light source to achieve uniform illumination, and the image sensor is used to image the magnetic beads in the collection area to obtain the concentration of the sample to be tested based on the fixed number of magnetic beads in the collection area.

[0018] In some embodiments, the light-docking area of ​​the light-docking plate is 2 to 5 times larger than the area of ​​the collection area;

[0019] And / or, the size of a single pixel of the image sensor is no larger than 700 nm;

[0020] And / or, the signal acquisition module further includes a cedar oil coating, and the cedar oil coating is located on the image sensor;

[0021] And / or, the signal acquisition module further includes a window protection sheet, and the window protection sheet is arranged on the image sensor.

[0022] The above-mentioned microfluidic single-molecule immunoassay analyzer uses microfluidic technology, uses less sample, has a fast detection speed, is simple to operate, and has a compact structure. It can integrate multifunctional mechanisms into one body and is small in size and easy to carry. The process of sample addition, dilution, reagent addition, mixing, incubation, reaction, cleaning, and signal acquisition are all completed on the microfluidic chip and corresponding mechanisms. The whole device is more compact, lower in cost, higher in sensitivity, lighter in weight, and faster in response. It can be designed as a micro-tabletop device, a handheld device, or a wearable device. It can be used in medical institutions of all levels and types, and can also be used by individual users at home, with a wide range of applications. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] To more clearly illustrate the technical solutions in the embodiments of the present application, the following briefly introduces the drawings required for describing the embodiments. Obviously, the drawings described below are only some embodiments of the present application. Those skilled in the art can also derive other drawings based on these drawings without inventive effort.

[0024] In order to more completely understand the present application and its beneficial effects, the following description will be given in conjunction with the accompanying drawings. In the following description, the same reference numerals represent the same parts.

[0025] Figure 1 Schematic diagram of a microfluidic chip of a microfluidic single molecule immunoassay analyzer according to one embodiment of the present invention;

[0026] Figure 2 This is a schematic diagram of a pressing module of a microfluidic single-molecule immunoassay analyzer according to an embodiment of the present invention;

[0027] Figure 3 This is a schematic diagram of the magnetic control module and collection area of ​​the microfluidic single-molecule immunoassay according to one embodiment of the present invention;

[0028] Figure 4 This is a schematic diagram of the working state of the magnetic control module and the first electromagnet of the collection area of ​​the microfluidic single-molecule immunoassay according to one embodiment of the present invention;

[0029] Figure 5 This is a schematic diagram of the working state of the magnetic control module and the second electromagnet in the collection area of ​​the microfluidic single-molecule immunoassay according to one embodiment of the present invention;

[0030] Figure 6 This is a schematic diagram of the magnetic control module and the air pressure pulse of the collection area of ​​the microfluidic single-molecule immunoassay according to one embodiment of the present invention;

[0031] Figure 7 This is a schematic diagram of a signal acquisition module of a microfluidic single-molecule immunoassay analyzer according to an embodiment of the present invention;

[0032] Figure 8 This is a schematic diagram of imaging of a signal acquisition module according to an embodiment of the present invention.

[0033] Description of Reference Numerals

[0034] 100. Microfluidic chip; 11. Test group; 12. Reference group; 101. 101a. Sample area; 102. 102a. Reagent area; 103. 103a. Collection area; 104. 104a. Waste liquid area; 105. 105a. First diluent area; 106. 106a. Second diluent area; 107. Test microchannel; 108. Reference microchannel; 200. Pressing module; 201. Driving component; 202. Pressing head; 300. Magnetron module; 301. First electromagnet; 302. Second electromagnet; 400. Signal acquisition module; 401. LED white light source; 402. Light homogenizer; 403. Image sensor; 404. Cedar oil coating; 405. Window protective layer; 20. Magnetic beads. DETAILED DESCRIPTION

[0035] To make the above-mentioned objects, features, and advantages of the present invention more clearly understood, the following detailed description of specific embodiments of the present invention is provided in conjunction with the accompanying drawings. The following description sets forth many specific details to facilitate a full understanding of the present invention. However, the present invention can be implemented in many other ways than those described herein, and those skilled in the art may make similar modifications without departing from the scope of the present invention. Therefore, the present invention is not limited to the specific embodiments disclosed below.

[0036] In the description of the present invention, it should be understood that the terms "center", "longitudinal", "lateral", "length", "width", "thickness", "up", "down", "front", "back", "left", "right", "vertical", "horizontal", "top", "bottom", "inside", "outside", "clockwise", "counterclockwise", "axial", "radial", "circumferential" and the like to indicate orientations or positional relationships based on the orientations or positional relationships shown in the accompanying drawings, and are only for the convenience of describing the present invention and simplifying the description, and do 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 should not be understood as a limitation to the present invention.

[0037] In this utility model, unless otherwise specified or limited, the terms "installed," "connected," "connect," "fixed," etc. should be understood in a broad sense. For example, they can refer to fixed connection, detachable connection, or integration; mechanical connection, electrical connection; direct connection, or indirect connection through an intermediate medium; internal communication between two components, or interaction between two components, unless otherwise specified. Those skilled in the art will understand the specific meanings of the above terms in this utility model based on specific circumstances.

[0038] In the present invention, unless otherwise expressly specified or limited, when a first feature is "above" or "below" a second feature, it may mean that the first and second features are in direct contact, or the first and second features are in indirect contact through an intermediary. Furthermore, when a first feature is "above," "above," or "above" a second feature, it may mean that the first feature is directly above or diagonally above the second feature, or simply means that the first feature is at a higher level than the second feature. When a first feature is "below," "below," or "below" a second feature, it may mean that the first feature is directly below or diagonally below the second feature, or simply means that the first feature is at a lower level than the second feature.

[0039] In the description of this utility model, "several" means more than one, "plurality" means more than two, "greater than," "less than," and "exceed" are understood to exclude the number itself, while "above," "below," and "within" are understood to include the number itself. The use of the terms "first" and "second" is solely for the purpose of distinguishing technical features and is not to be construed as indicating or implying relative importance, implicitly specifying the number of the indicated technical features, or implicitly specifying the order of the indicated technical features.

[0040] Unless otherwise defined, all technical and scientific terms used herein have the same meanings as those commonly understood by those skilled in the art to which this invention pertains. The terms used herein in the specification of this invention are for the purpose of describing specific embodiments only and are not intended to limit this invention. The term "and / or" as used herein includes any and all combinations of one or more of the associated listed items.

[0041] In this application, when referring to a numerical interval (i.e., a numerical range), unless otherwise specified, the distribution of the optional numerical values ​​within the numerical interval is deemed to be continuous and includes the two numerical endpoints of the numerical interval (i.e., the minimum and maximum values), as well as each numerical value between the two numerical endpoints. Unless otherwise specified, when a numerical interval refers only to integers within the numerical interval, it includes the two endpoint integers of the numerical range, as well as each integer between the two endpoints, which is equivalent to directly listing each integer. When multiple numerical ranges are provided to describe a feature or characteristic, these numerical ranges can be combined. In other words, unless otherwise specified, the numerical ranges disclosed in this application should be understood to include any and all subranges included therein. The "numerical value" in the numerical interval can be any quantitative value, such as a number, percentage, ratio, etc. "Numerical interval" is broadly allowed to include quantitative intervals such as percentage intervals, ratio intervals, and ratio intervals.

[0042] The present invention provides a microfluidic single-molecule immunoassay analyzer to address the challenges of conventional immunoassay detection instruments, such as complex overall structure, bulk, high cost, difficult microarray processing, and the need for external excitation light paths with various wavelengths for fluorescence signals, resulting in a complex structure. The microfluidic single-molecule immunoassay analyzer is described below with reference to the accompanying figures.

[0043] The microfluidic single molecule immunoassay analyzer provided in the present application is exemplary, see Figure 1 As shown, Figure 1 Schematic diagram of the microfluidic chip 100 of the microfluidic single-molecule immunoassay analyzer provided in an embodiment of the present application. The microfluidic single-molecule immunoassay analyzer of the present application can be used for immunoassay purposes. The present application can be combined with the double antibody sandwich method for detection.

[0044] In order to more clearly illustrate the structure of the microfluidic single molecule immunoassay analyzer, the microfluidic single molecule immunoassay analyzer will be introduced below with reference to the accompanying drawings.

[0045] For example, a microfluidic single molecule immunoassay analyzer includes a microfluidic chip 100, a motion control module, a temperature control module, a pressing module 200, a magnetic control module 300, and a signal acquisition module 400. Figure 1 As shown, the microfluidic chip 100 includes a sample area 101, a reagent area 102, a collection area 103, and a waste liquid area 104. The motion control module and the temperature control module are not shown in the drawings.

[0046] See also Figure 1As shown, the sample area 101 and the reagent area 102 are connected to the collection area 103 through the test microchannel 107. The collection area 103 is connected to the waste liquid area 104. The sample area 101 is used to add a certain amount of sample to be tested. The reagent area 102 contains magnetic beads 20 coated with antibodies. The collection area 103 is connected with antibodies and can adsorb the magnetic beads 20. The waste liquid area 104 is used to collect free magnetic beads 20 that are not adsorbed. The motion control module is used to drive the microfluidic chip 100 to pass through the pressing module 200, the magnetic control module 300 and the signal acquisition module 400 in sequence. The pressing module 200 is used to realize the addition of the first diluent into the sample area 101 to realize the mixed incubation reaction of the sample to be tested, the whole blood treatment reagent and the first diluent in the sample area 101 to form the antigen to be tested. The pressing module 200 is also used to add a second diluent to the reagent zone 102 to dissolve the freeze-dried antibody-coated magnetic beads 20 in the reagent zone 102 into a liquid state. The magnetic control module 300 is used to ensure that the antibodies on the magnetic beads 20 bind to the antigen to be tested and are adsorbed into the collection zone 103. The collection module is used to detect the number of magnetic beads 20 in the collection zone 103, and the temperature control module is used to control the test temperature.

[0047] The above-mentioned microfluidic single-molecule immunoassay analyzer uses microfluidic technology, uses less sample, has a fast detection speed, is simple to operate, and has a compact structure. It can integrate multifunctional mechanisms into one body and is small and easy to carry. The process of sample addition, dilution, reagent addition, mixing, incubation, reaction, cleaning, and signal acquisition are all completed on the microfluidic chip 100 and its corresponding mechanisms. The whole device is more compact, lower in cost, higher in sensitivity, lighter in weight, and has a fast response speed. It can be designed as a micro-tabletop device, a handheld device, or a wearable device. It can be used in various medical institutions at all levels and can also be used by individual users at home, with a wide range of applications.

[0048] In some embodiments, the reagent area 102 contains magnetic beads 20 coated with antibodies, which can be in the form of both lyophilized reagents and liquid reagents, or in the form of only lyophilized reagents or only liquid reagents.

[0049] In some embodiments, the sample area 101 is pre-loaded with whole blood processing reagents.

[0050] In some of these examples, see Figure 1 As shown, the microfluidic chip 100 further includes a first diluent area 105. The first diluent area 105 is connected to the sample area 101 to add a first diluent into the sample area 101, thereby incubating the sample to be tested, the whole blood treatment reagent, and the first diluent in the sample area 101 to form the antigen to be tested.

[0051] In some of these examples, see Figure 1As shown, the microfluidic chip 100 further includes a second diluent area 106. The second diluent area 106 is connected to the reagent area 102 to add a second diluent into the reagent area 102, so that the lyophilized antibody-coated magnetic beads 20 in the reagent area 102 are dissolved into liquid.

[0052] In some of these examples, see Figure 1 As shown, the microfluidic chip 100 includes at least one test group 11 and at least one reference group 12. The test group 11 and the reference group 12 both include a sample area 101, a reagent area 102, a collection area 103, and a waste liquid area 104. The sample area 101a and the reagent area 102a in the reference group 12 are connected to the corresponding collection area 103a through a reference microchannel 108. The reference microchannel 108 is the same length as the test microchannel 107. The sample area 101a in the reference group 12 contains an artificially synthesized antigen to be tested. The first diluent and the second diluent in the test group 11 and the reference group 12 are added synchronously through the same pressing module 200.

[0053] Furthermore, test group 11 includes a sample area 101, a reagent area 102, a collection area 103, a waste liquid area 104, a first diluent area 105, and a second diluent area 106. Reference group 12 includes a sample area 101a, a reagent area 102a, a collection area 103a, a waste liquid area 104a, a first diluent area 105a, and a second diluent area 106a. In other words, the structures of test group 11 and reference group 12 are essentially the same, with one group serving as a test and the other as a control.

[0054] Preferably, see Figure 1 As shown, a test group 11 and a reference group 12 are simultaneously provided on a microfluidic chip 100. The test group 11 and the reference group 12 can be added with the first diluent and the second diluent through the same pressing module 200.

[0055] It should be noted that the reference group 12 can also use the structures and functions of the following test group 11 to perform corresponding operation steps. The following description will be based on the test group 11.

[0056] In some of these examples, see Figure 2As shown, the pressing module 200 includes a driving component 201 and at least two pressing heads 202. The driving component 201 is connected to the pressing heads 202 to drive the pressing heads 202 to press synchronously. When the motion control module drives the microfluidic chip 100 to move to the first diluent area 105 located at the pressing module 200, the driving component 201 drives the pressing heads 202 to press synchronously to add the first diluent in the first diluent area 105 to the sample area 101. When the motion control module drives the microfluidic chip 100 to move to the second diluent area 106 located at the pressing module 200, the driving component 201 drives the pressing heads 202 to press synchronously to add the second diluent in the second diluent area 106 to the reagent area 102.

[0057] In some embodiments, the driving component 201 of the pressing module 200 can be a pressing pusher or a pneumatic pusher. The pressing module 200 can add samples to the test group 11 and the reference group 12 at the same time.

[0058] In some of these examples, see Figure 3 As shown, the magnetic control module 300 includes a first electromagnet 301 and a second electromagnet 302. The first electromagnet 301 and the second electromagnet 302 are spaced apart in the vertical direction. The motion control module can drive the microfluidic chip 100 to move between the first electromagnet 301 and the second electromagnet 302. The magnetic poles of the first electromagnet 301 and the second electromagnet 302 can alternate.

[0059] When the collection area 103 of the microfluidic chip 100 is located on the magnetic control module 300, the first electromagnet 301 and the second electromagnet 302 alternate in a certain waveform relationship, accelerating the movement of the antibody-labeled magnetic beads 20 in the liquid, increasing the probability of binding with the antigen to be tested. This changes the binding mode from pure Brownian diffusion to a binding mode with an external driving force, reducing incubation time and improving detection efficiency. The magnetic control module 300 can simultaneously magnetically control the test group 11 and the reference group 12.

[0060] See also Figure 4 As shown, depending on the concentration of the antigen to be tested, some magnetic beads bind to the antigen to be tested, while some magnetic beads do not. When the first electromagnet 301 located below is working, the first electromagnet 301 attracts the magnetic beads 20 and accelerates them downward. Under the adsorption of the lower magnet, all magnetic beads bound to the antigen to be tested and magnetic beads not bound to the antigen to be tested are sucked down and move toward the bottom. The magnetic beads 20 bound to the antigen to be tested and coated with antibodies bind to the antibodies coated on the surface of the chip to form a double antibody sandwich. The magnetic beads 20 that are not bound to the antigen to be tested and coated with antibodies are magnetically attracted downward. The magnetic beads that are not bound to the antigen to be tested cannot bind to the antibodies coated on the substrate and cannot form a double antibody sandwich; please refer to Figure 5As shown, when the second electromagnet 302 located above is activated, the magnetic force exerted by the second electromagnet 302 is greater than the gravity of the magnetic beads 20. The adsorption force of the magnetic beads forming the double antibody sandwich is less than the gravity plus the antigen-antibody binding force, preventing them from moving upward. The magnetic beads not forming the double antibody sandwich experience an adsorption force greater than the gravity, moving upward and away from the bottom of the chip. Consequently, the antibody-coated magnetic beads 20 not fixed to the chip are pulled upward and away from the bottom of the chip, entering the collection area 103.

[0061] In some embodiments, the magnetic control module 300 further includes an air pressure pulse component (not shown in the drawings). The air pressure pulse component is used to apply air pressure pulses to the collection area 103. When the first electromagnet 301 located below is working, the collection area 103 is driven by the air pressure pulse to move the magnetic beads 20 back and forth, so that they can be evenly distributed in the collection area 103, reducing aggregation or stacking. Figure 6 As shown, when the second electromagnet 302 located above is working, the collection area 103 is driven by the air pressure pulse to accelerate the magnetic beads 20 to separate from the collection area 103.

[0062] In some embodiments, the magnetic pole area of ​​the first electromagnet 301 is 2 to 5 times larger than the area of ​​the collection region 103 .

[0063] In some embodiments, the magnetic pole area of ​​the second electromagnet 302 is 2 to 5 times larger than the area of ​​the collection area 103.

[0064] In some of these examples, see Figure 7 As shown, the signal acquisition module 400 includes an LED white light source 401, a light diffuser 402, and an image sensor 403. The light diffuser 402 and the image sensor 403 are spaced apart in the vertical direction, and the microfluidic chip 100 can pass between the light diffuser 402 and the image sensor 403. The LED white light source 401 is used to provide brightfield imaging illumination, and the light diffuser 402 is used to diffuse the LED white light source 401 to achieve uniform illumination. The image sensor 403 is used to image the magnetic beads 20 in the collection area 103 to obtain the concentration of the sample to be tested based on the number of magnetic beads 20 fixed in the collection area 103. The image sensor 403 directly images the magnetic beads 20, obtains the number of magnetic beads 20 fixed in the collection area 103, and determines the concentration value of the sample to be tested. The minimum detection limit of the image sensor 403 is one magnetic bead 20, and the upper limit is the total number of magnetic beads 20 that can be input.

[0065] In some embodiments, the light-diffusing area of ​​the light-diffusing plate 402 is 2 to 5 times larger than the area of ​​the collection area 103 .

[0066] It should be noted that the magnetic pole area of ​​the first electromagnet 301, the magnetic pole area of ​​the second electromagnet 302, and the uniform light area of ​​the light diffuser 402 are respectively 2 to 5 times the area of ​​the collection area 103. This is an experimental value and can be adjusted according to actual effects, chips, circuit capabilities and other factors. However, the magnetic pole area of ​​the first electromagnet 301, the magnetic pole area of ​​the second electromagnet 302, and the uniform light area of ​​the light diffuser 402 must all be larger than the area of ​​the collection area 103.

[0067] In some embodiments, the image sensor 403 may be a CMOS image sensor 403 or a CCD image sensor 403. The image sensor 403 may be an area array image sensor 403 or a line array image sensor 403.

[0068] In some embodiments, the CMOS image sensor 403 images the Figure 8 As shown, the size of a single pixel of the CMOS image sensor 403 imaging is no more than 700nm. The key point of this application lies in the mutual constraints between the size of the magnetic beads 20, the performance of the reagent, the CMOS pixel size and the total size. An optimized combination is that the particle size of the magnetic beads 20 is 3μm, the imaging is clear and the number of magnetic beads 20 can be distinguished, and the size of a single pixel of the image sensor 403 imaging needs to be no more than 700nm to avoid the superposition of magnetic beads 20 and magnetic beads 20 affecting the imaging. The spacing between magnetic beads 20 and magnetic beads 20 must be maintained at no less than 2μm, and the actual size occupied by a single magnetic bead 20 is about 5μm. Then the measurable range of the sample to be tested is equal to the maximum number of magnetic beads 20 that can be distributed. For example, when the total size of the image sensor 403 is 5mm×5mm, the number of magnetic beads 20 that can be distributed is 1000×1000, a total of 1 million, and the total number of pixels required for the CMOS is about 52 million. The 5mm×5mm size of acquisition area 103 is only an example and needs to be modified to fit the chip size of CMOS image sensor 403, so that it can be smaller than the chip size of CMOS image sensor 403. However, the size of magnetic beads 20 also determines the maximum testable range. For example, if 1μm magnetic beads 20 are used, the maximum testable range within acquisition area 103 is 5000×5000=25 million.

[0069] In some embodiments, the signal acquisition module 400 further includes a cedar oil coating 404 . The cedar oil coating 404 is located on the image sensor 403 .

[0070] In some embodiments, the signal acquisition module 400 further includes a window protection sheet. The window protection sheet is positioned over the image sensor 403 to prevent damage to the image sensor 403. The space between the image sensor 403 and the window protection sheet is filled with colorless, transparent cedar oil. The cedar oil coating 404 allows more light to enter the image sensor 403, resulting in a brighter field of view and improved image clarity.

[0071] In some embodiments, the overall shape of the microfluidic chip 100 of the present application can be a rectangular design or other shapes.

[0072] In some embodiments, the temperature of the temperature control module is controlled to 37° C. The optimal temperature of 37° C. is the temperature used in most detection methods, but it can also be changed to other temperatures according to actual needs to meet the test requirements.

[0073] In summary, compared with traditional technologies, this application has the following beneficial effects:

[0074] (1) This application adopts an alternating electromagnetic field control method to achieve drive control of each process of the immune response.

[0075] (2) This application adopts the pressing push of the pressing module 200, combined with electromagnetic field control and motor control, so that the magnetic beads 20 are evenly distributed in the collection area 103, avoiding the magnetic beads 20 from gathering together and affecting the final result.

[0076] (3) This application uses a lensless CMOS image sensor 403 for image acquisition. Due to the use of a lensless image acquisition method, the total volume of the signal acquisition module 400 is greatly reduced, the focusing system is eliminated, and the control complexity is greatly reduced.

[0077] (4) This application adopts a large-area array CMOS image sensor 403, which can complete the reading of all data of a project in one acquisition, and the acquisition speed is fast.

[0078] (5) The microfluidic chip 100 does not require microarray micropore processing, which reduces the production complexity and single-chip production and processing costs. The magnetic beads 20 are distributed on the plane and will not sink into the small holes. When the microfluidic chip 100 is cleaned after the test, the magnetic beads 20 are more easily washed away.

[0079] (6) During the test, the CMOS image sensor 403 is located below the microfluidic chip 100. The CMOS image sensor 403 takes photos upward, avoiding the influence of bubbles on the surface of the liquid in the collection area 103. At the same time, since the liquid enclosed in the microfluidic chip 100 will generate steam when heated, the CMOS image sensor 403 takes photos upward to avoid condensation on the surface of the microfluidic chip 100 that affects the photographic effect.

[0080] (7) A reference group 12 is set up. The sample area 101 in the reference group 12 contains artificial antigens and represents the maximum test range. The antibody-coated magnetic beads 20 and the microfluidic chip 100 are from the same batch to ensure consistency. The test content is calculated by comparing and normalizing the images of the test group 11 of the test microchannel 107 and the reference group 12 of the reference microchannel 108. Therefore, the present application does not require micromatrix processing on the microfluidic chip 100, reducing chip costs.

[0081] In the above embodiments, the description of each embodiment has its own focus. For parts that are not described in detail in a certain embodiment, reference can be made to the relevant descriptions of other embodiments.

[0082] The technical features of the above-mentioned embodiments can be combined arbitrarily. In order to make the description concise, not all possible combinations of the technical features in the above-mentioned 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.

[0083] The above-described embodiments merely represent several implementation methods of the present invention. While the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the present invention. It should be noted that a person skilled in the art would be able to make various modifications and improvements without departing from the concept of the present invention, and these modifications and improvements fall within the scope of protection of the present invention. Therefore, the scope of protection of the present invention shall be determined by the appended claims.

Claims

1. A microfluidic single molecule immunoassay analyzer, characterized in that: The invention comprises a microfluidic chip, a motion control module, a temperature control module, a pressing module, a magnetic control module and a signal acquisition module, wherein the microfluidic chip comprises a sample area, a reagent area, a collection area and a waste liquid area, the sample area and the reagent area are connected to the collection area through a test microchannel, the collection area is connected to the waste liquid area, the sample area is used to add a certain amount of sample to be tested, the reagent area contains magnetic beads coated with antibodies, the collection area is connected with antibodies and can achieve adsorption of magnetic beads, the waste liquid area is used to collect free magnetic beads that are not adsorbed, and the motion control module is used to drive the microfluidic chip to pass through the test microchannel in sequence. The pressing module, the magnetic control module and the signal acquisition module, the pressing module is used to add the first diluent into the sample area to realize a mixed incubation reaction of the sample to be tested, the whole blood processing reagent and the first diluent in the sample area to form the antigen to be tested, the pressing module is also used to realize the addition of the second diluent into the reagent area to realize the dissolution of the freeze-dried antibody-coated magnetic beads in the reagent area into a liquid state, the magnetic control module is used to realize the antibodies on the magnetic beads being adsorbed into the acquisition area by binding to the antigen to be tested, the acquisition module is used to detect the number of magnetic beads in the acquisition area, and the temperature control module is used to control the test temperature.

2. The microfluidic single molecule immunoassay analyzer according to claim 1, characterized in that: The sample area is pre-installed with whole blood processing reagents.

3. The microfluidic single molecule immunoassay analyzer according to claim 1, characterized in that: The microfluidic chip also includes a first diluent area, which is connected to the sample area to enable the first diluent to be added into the sample area, so as to achieve a mixed incubation reaction of the test sample, the whole blood processing reagent and the first diluent in the sample area to form the test antigen.

4. The microfluidic single molecule immunoassay analyzer according to claim 1, characterized in that: The microfluidic chip further includes a second diluent area, which is connected to the reagent area to enable the second diluent to be added into the reagent area, thereby dissolving the freeze-dried antibody-coated magnetic beads in the reagent area into a liquid state.

5. The microfluidic single molecule immunoassay analyzer according to any one of claims 1 to 4, characterized in that: The microfluidic chip includes at least one test group and at least one reference group. The test group and the reference group both include the sample area, the reagent area, the collection area and the waste liquid area. The sample area and the reagent area in the reference group are connected to the corresponding collection area through a reference microchannel. The reference microchannel and the test microchannel have the same length. The sample area in the reference group contains an artificially synthesized antigen to be tested. The first diluent and the second diluent in the test group and the reference group are synchronously added through the same pressing module.

6. The microfluidic single molecule immunoassay analyzer according to claim 5, characterized in that: The pressing module includes a driving component and at least two pressing heads. The driving component is connected to the pressing head to drive the pressing head to press synchronously. When the motion control module drives the microfluidic chip to move to the first diluent area located at the pressing module, the driving component drives the pressing head to press synchronously to add the first diluent in the first diluent area to the sample area; when the motion control module drives the microfluidic chip to move to the second diluent area located at the pressing module, the driving component drives the pressing head to press synchronously to add the second diluent in the second diluent area to the reagent area.

7. The microfluidic single-molecule immunoassay analyzer according to any one of claims 1 to 4 and 6, characterized in that: The magnetic control module includes a first electromagnet and a second electromagnet, the first electromagnet and the second electromagnet are spaced apart in the vertical direction, the motion control module is capable of driving the microfluidic chip to move between the first electromagnet and the second electromagnet, and the magnetic poles of the first electromagnet and the second electromagnet are capable of alternating; When the first electromagnet located at the bottom is working, it attracts the magnetic beads and accelerates them to move downward. The magnetic beads bound to the antigen bind to the antibodies coated on the surface of the chip, and the magnetic beads not bound to the antigen are magnetically sucked downward; when the second electromagnet located at the top is working, the magnetic force of the second electromagnet is controlled to be greater than the gravity of the magnetic beads, and the magnetic beads not fixed on the chip are sucked upward and separated from the bottom of the chip and enter the collection area.

8. The microfluidic single molecule immunoassay analyzer according to claim 7, characterized in that: The magnetic control module further includes an air pressure pulse component for applying an air pressure pulse to the collection area. When the first electromagnet located below is in operation, the collection area is driven by the air pressure pulse to move the magnetic beads back and forth, so that they are evenly distributed in the collection area and reduce aggregation or stacking. When the second electromagnet located above is in operation, the collection area is driven by the air pressure pulse to accelerate the magnetic beads out of the collection area. And / or, the magnetic pole area of ​​the first electromagnet is 2 to 5 times larger than the area of ​​the collection area; And / or, the magnetic pole area of ​​the second electromagnet is 2 to 5 times larger than the area of ​​the collection area.

9. The microfluidic single molecule immunoassay analyzer according to any one of claims 1 to 4, 6, and 8, characterized in that: The signal acquisition module includes an LED white light source, a light-diffusing plate, and an image sensor; The light homogenizing plate and the image sensor are spaced apart in the vertical direction, and the microfluidic chip can pass between the light homogenizing plate and the image sensor. The LED white light source is used to provide bright field imaging illumination, and the light homogenizing plate is used to homogenize the LED white light source to achieve uniform illumination. The image sensor is used to image the magnetic beads in the collection area to obtain the concentration of the sample to be tested based on the fixed number of magnetic beads in the collection area.

10. The microfluidic single molecule immunoassay analyzer according to claim 9, characterized in that: The light homogenizing area of ​​the light homogenizing plate is 2 to 5 times larger than the area of ​​the collection area; And / or, the size of a single pixel of the image sensor is no larger than 700 nm; And / or, the signal acquisition module further includes a cedar oil coating, and the cedar oil coating is located on the image sensor; And / or, the signal acquisition module further includes a window protection sheet, and the window protection sheet is arranged on the image sensor.

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