Microfluidic magnetic microparticle chemiluminescence reaction device

Abstract

The present invention discloses a microfluidic magnetic particle chemiluminescence reaction device, which belongs to the field of medical inspection technology. The device includes a sample adding block, a capillary glass tube, a hydrophobic ring, a hydrophobic disk, a rotary drive mechanism and an electromagnet; the sample adding block is provided with a sample adding hole, one end of the capillary glass tube is sealed and connected with the sample adding block and is connected with the sample adding hole; the hydrophobic ring is arranged on the outer peripheral surface of the end of the capillary glass tube away from the sample adding block, which is used to enhance the surface tension of the reaction liquid flowing thereto; the hydrophobic disk is connected to the rotary drive mechanism, which is used to break the surface tension of the reaction liquid and throw the reaction liquid out from the end of the capillary glass tube; the electromagnet is arranged on the periphery of the capillary glass tube, which is used to disperse and position the magnetic particles. The present application uses a microfluidic pump composed of a hydrophobic ring, a hydrophobic disk and a rotary drive mechanism to control the flow of the liquid. The liquid flow speed is relatively slow and the reaction process is controllable, which can achieve high sensitivity and accurate quantitative measurement of the target substance.

Description

Technical Field

[0001] The invention relates to the technical field of medical testing, and in particular to a microfluidic magnetic particle chemiluminescence reaction device. Background Art

[0002] The chemiluminescent immunoassay technology has developed into a mature and advanced technology for detecting ultra-trace active substances. Its sensitivity and accuracy are higher than those of enzyme immunoassay and fluorescence method, and its application range is very wide.

[0003] Chemiluminescent immunoassay includes two parts, namely, immunoreaction technology and chemiluminescence technology. The basic principle is that the enzyme in the immunoreaction acts on the luminescent substrate, causing it to undergo a chemical reaction and release a large amount of energy, producing an excited intermediate. When this excited intermediate returns to a stable ground state, it can emit photons at the same time. The light quantum yield can be measured using a luminescent signal measuring instrument. The light quantum yield is proportional to the amount of the substance to be tested in the sample, so a standard curve can be established and the content of the substance to be tested in the sample can be calculated.

[0004] Magnetic particle chemiluminescence immunoassay technology combines magnetic particle carrier technology and chemiluminescence immunoassay technology. Magnetic particles refer to colloidal composite materials formed by the combination of magnetic nanoparticles and inorganic or organic molecules that can be evenly dispersed in a certain base liquid and have high stability. There are three main methods for magnetic particle chemiluminescence reaction: double antibody sandwich method, competitive method and indirect method. Based on the characteristics of magnetic particles such as magnetic responsiveness, low cost, low energy consumption and no pollution, enzymes, antibodies and other bioactive substances are fixed on the surface of magnetic particles, which can be further used in enzyme immobilization, cell sorting, immunoassay and other fields.

[0005] Traditional microfluidic magnetic particle chemiluminescence reaction devices mainly rely on the power pump at the end of the device to suck away the liquid. The pumping equipment has the characteristics of strong power and rapid reaction process. However, based on this characteristic of the pumping equipment, the flow rate of the reaction liquid is poorly controllable, and it is easy to quickly extract the reaction liquid from the reaction tube in one go without fully reacting. The detection sensitivity and accuracy cannot be guaranteed; and most of the existing reaction devices use pipes carved by engraving machines as the main reaction containers. The inner wall of the pipe is not smooth enough, and the reaction liquid sticks to the wall obviously, which is difficult to clean thoroughly, which will interfere with the accuracy of the test results.

[0006] Chinese patent CN107478837A also discloses a microfluidic chemiluminescence detection system based on magnetic particles that controls the reaction process by centrifugation, comprising a chassis and an upper chip; a sample loading area and at least one microfluidic reaction detection channel connected to the sample loading area are provided on the upper chip; the microfluidic reaction detection channel comprises: a sample separation area that is connected to the sample loading area; an antibody coating area that is connected to the sample separation area through a capillary glass microchannel; a magnetic particle coating area that is connected to the antibody coating area through a capillary glass microchannel; a waste liquid collection area that is connected to the magnetic particle coating area through a capillary glass microchannel; the antibody coating area is coated with an antibody to a test object labeled with a luminescent substance and another antibody to a test object labeled with one of a pair of substances with specific affinity; the magnetic particle coating area is coated with magnetic particles labeled with the other of the pair of substances with specific affinity; the chassis is arranged below the upper chip, and a magnet is arranged at a position of the chassis corresponding to the magnetic particle coating area. It mainly uses centrifugal action and magnetic field action to sequentially and orderly realize the preparation of immune complexes and magnetic microparticle immune complexes, collection of waste liquid, washing of immune complexes, and addition of luminescent substrate liquid, and finally realizes quantitative measurement of the object to be tested. Although the system can realize rapid and high-sensitivity measurement of the target substance, it still has the problem of poor controllability of reaction speed when relying on centrifugal action to complete each step of the reaction. The reaction liquid is easily thrown out quickly during the centrifugation process, and the stability of the sequential flow is poor. It is difficult to realize the intermittent and orderly flow of the reaction liquid from the sample addition area-sample separation area-antibody coating area-magnetic microparticle coating area-waste liquid collection area. It cannot ensure that each step of the reaction is carried out in an orderly and sufficient manner, and the controllability of the reaction process still needs to be improved. Summary of the invention

[0007] The purpose of the present invention is to solve the problem of insufficient controllability of traditional microfluidic magnetic particle chemical reaction devices in which the reaction process is controlled by a pumping device, and to provide a new microfluidic magnetic particle chemiluminescence reaction device, which uses a microfluidic pump composed of a hydrophobic ring, a hydrophobic disk and a rotating drive mechanism to control the flow of liquid. The liquid flow speed is relatively slow and the reaction process is controllable, which can achieve high-sensitivity and accurate quantitative measurement of the target substance.

[0008] The technical scheme disclosed in the present invention is: a microfluidic magnetic particle chemiluminescence reaction device, comprising a sample adding block, a capillary glass tube, a hydrophobic ring, a hydrophobic disk, a rotating drive mechanism and an electromagnet; the sample adding block is provided with a sample adding hole for dripping a reaction liquid; the capillary glass tube uses capillary action to guide the reaction liquid, one end of which is sealed and connected with the sample adding block and is connected with the sample adding hole; the hydrophobic ring is used to enhance the surface tension of the reaction liquid, and the hydrophobic ring is arranged on the outer peripheral surface of the end of the capillary glass tube away from the sample adding block; the hydrophobic disk is used to break the surface tension of the reaction liquid, and the hydrophobic disk is connected with the rotating drive mechanism; the electromagnet is used to disperse and position the magnetic particles, and the electromagnet is arranged on the periphery of the capillary glass tube.

[0009] Furthermore, the hydrophobic disk is arranged in parallel with the hydrophobic ring, and the distance between the hydrophobic ring and the hydrophobic disk is 0.01-0.50 mm.

[0010] Furthermore, the hydrophobic disk is eccentrically arranged relative to the hydrophobic ring, and both the hydrophobic ring and the hydrophobic disk are made of hydrophobic material.

[0011] Furthermore, three electromagnets are arranged outside the capillary glass tube, namely electromagnet one, electromagnet two and electromagnet three.

[0012] Furthermore, electromagnet one and electromagnet two are respectively arranged on the front and rear sides of the capillary glass tube and are coaxially symmetrically arranged, and electromagnet three is arranged directly below the capillary glass tube, and the axis of electromagnet three is perpendicular to the axis of electromagnet one.

[0013] Furthermore, the optical signal detection device is arranged above the capillary glass tube and corresponds to the position of the electromagnet.

[0014] Furthermore, the capillary glass tube is made of a highly light-transmitting hydrophilic material.

[0015] Furthermore, the rotation driving mechanism is a motor, and the side of the hydrophobic disk away from the hydrophobic ring is coaxially fixedly connected to the motor shaft of the motor.

[0016] The beneficial effects of the present invention are:

[0017] 1. The present application uses a microfluidic pump composed of a hydrophobic ring, a hydrophobic disk and a rotating drive mechanism to control the outflow process of the liquid in the capillary glass tube. When the hydrophobic disk is stationary, the liquid flows from the sample addition end to the hydrophobic ring end under the capillary action, and then stops flowing due to the hydrophobic effect and forms an outwardly convex liquid film under the action of surface tension. When the motor drives the hydrophobic disk to rotate, the hydrophobic disk will destroy the surface tension of the liquid, and the molecules on the surface of the hydrophobic disk drive the liquid to rotate, and under the action of centrifugal force, the liquid at the end of the capillary glass tube is continuously thrown out, and then the liquid in the capillary glass tube can be gradually discharged. Compared with traditional high-power pump suction equipment or devices that use centrifugal action to control liquid flow, this new type of microfluidic pump structure uses hydrophobic action to achieve liquid discharge, which can slow down the liquid flow rate, ensure that each step of the reaction is fully carried out, and will not cause the liquid to be quickly discharged from the port when the reaction is incomplete, which helps to improve the detection sensitivity and accuracy;

[0018] 2. The present application uses a microfluidic pump structure to slow down the flow rate of the reaction liquid. Since the hydrophobic disk is controlled to rotate by a motor, the flow rate of the liquid can be further regulated by adjusting the speed of the motor. When the flow rate of the liquid is slowed down, the flow rate of the liquid is still in a state of artificial control, so that the detection efficiency and accuracy are further improved;

[0019] 3. The present application uses a capillary glass tube as the main reaction pipeline, and can use capillary action to achieve the purpose of self-running and continuous reaction. Compared with the reaction container engraved by an engraving machine, the capillary glass tube has a smooth tube wall, and it is not easy for the reaction liquid to stick to the wall, so that the process of washing the immune complex bound to the magnetic particles with a cleaning solution is smoother, and sensitive and accurate detection of the target substance can be achieved;

[0020] 4. The present application arranges electromagnets at the front, back and bottom of the capillary glass tube. The magnetism of the electromagnets is controllable, and the design of evenly distributed electromagnets at three angles makes it possible to balance the magnetic forces in three directions after power-on control, thereby achieving uniform dispersion of magnetic beads and effectively capturing low-concentration samples to be tested. Combined with the chemiluminescence detection method, the reaction sensitivity is greatly improved, ensuring that the reaction is fully carried out, and ultimately improving the accuracy and effectiveness of the test results;

[0021] 5. The magnetic microparticle chemiluminescence reaction device disclosed in the present application integrates the respective advantages of microfluidics, magnetic microparticles, and chemiluminescence detection methods. Microfluidics has the characteristics of miniaturization, microquantity, and high-throughput detection. Magnetic microparticles are easy to separate and clean, and chemiluminescence detection has high sensitivity. Therefore, the whole device can achieve rapid, high-sensitivity and accurate quantitative detection of target substances. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 It is a schematic diagram of the main view of a microfluidic magnetic particle chemiluminescence reaction device;

[0023] Figure 2 A schematic top view of a microfluidic magnetic particle chemiluminescence reaction device;

[0024] Figure 3 It is a right side schematic diagram of a microfluidic magnetic particle chemiluminescence reaction device;

[0025] Figure 4 for Figure 1 Cross-sectional view along direction BB;

[0026] Figure 5 for Figure 2 Cross-sectional view along AA direction;

[0027] Figure 6 It is an axonometric diagram of a microfluidic magnetic particle chemiluminescence reaction device;

[0028] Figure 7 It is a top view schematic diagram of a microfluidic magnetic particle chemiluminescence reaction device with two sets of detection pathways;

[0029] Among them, 1-sample block, 2-capillary glass tube, 3-hydrophobic ring, 4-hydrophobic disk, 5-motor, 6-electromagnet;

[0030] 11- sample well;

[0031] 61-electromagnet one, 62-electromagnet two, 63-electromagnet three. DETAILED DESCRIPTION

[0032] The following examples further illustrate the content of the present invention, but should not be construed as limiting the present invention. Without departing from the essence of the present invention, modifications and substitutions made to the methods, steps or conditions of the present invention all fall within the scope of the present invention. Example 1

[0033] In order to better control the flow process of liquid during the magnetic particle chemiluminescence reaction, ensure that each step of the reaction can be carried out fully and effectively, thereby achieving high-sensitivity detection of the target substance, and ultimately effectively improving the accuracy of the detection results, a microfluidic magnetic particle chemiluminescence reaction device is disclosed in this embodiment.

[0034] The device comprises a sample adding block 1, a capillary glass tube 2, a hydrophobic ring 3, a hydrophobic disk 4, a motor 5 and an electromagnet 6. A sample adding hole 11 is arranged on the sample adding block 1. The capillary glass tube 2 is made of a highly light-transmitting hydrophilic material. One end of the capillary glass tube 2 is sealedly connected to the sample adding block 1 and is connected to the sample adding hole 11 on the sample adding block 1. Of course, the capillary glass tube 2 can also be integrally processed and formed with the sample adding block 1, and the connection form is not specifically limited. The hydrophobic ring 3 is fixedly connected to the outer peripheral surface of the other end of the capillary glass tube 2. The side of the hydrophobic ring 3 away from the sample adding block 1 is flush with the end face of the capillary glass tube 2 away from the sample adding block 1. The liquid dripped into the sample adding block 1 can flow from the sample adding hole 11 along the capillary glass tube 2 toward the hydrophobic ring 3 based on the capillary action.

[0035] The hydrophobic disc 4 is arranged parallel to the hydrophobic ring 3 , and the side of the hydrophobic disc 4 away from the hydrophobic ring 3 is coaxially fixedly connected to the motor shaft of the motor 5 , and the distance between the hydrophobic ring 3 and the hydrophobic disc 4 is 0.01-0.50 mm.

[0036] The hydrophobic ring 3 and the hydrophobic disk 4 are both made of hydrophobic material. Preferably, the hydrophobic ring 3 and the hydrophobic disk 4 can be made of Teflon material.

[0037] The electromagnet 6 is arranged at the periphery of the capillary glass tube 2 and is biased toward one side of the hydrophobic ring 3. Under the action of the alternating current, the electromagnet 6 disperses and positions the magnetic particles (magnetic beads) in the liquid in the capillary glass tube 2. In order to better balance the magnetic force, so that the magnetic particles can be dispersed more evenly in the tube, conveniently and effectively capture the low-concentration samples to be tested in the samples to be tested, and greatly improve the reaction sensitivity after combining the chemiluminescence detection method, in this embodiment, three electromagnets 6 are arranged at the periphery of the capillary glass tube 2, namely, electromagnet 1 61, electromagnet 2 62 and electromagnet 3 63. Electromagnet 1 61 and electromagnet 2 62 are respectively arranged at the front and rear sides of the capillary glass tube 2 and are symmetrically arranged. Electromagnet 3 63 is arranged directly below the capillary glass tube 2. Electromagnet 1 61 and electromagnet 2 62 are coaxially arranged, and the axis of electromagnet 3 63 is perpendicular to the axis of electromagnet 1 61. The electromagnet 1 61 and the electromagnet 2 62 are driven by alternating current to disperse and position the magnetic particles in the liquid flowing in the capillary glass tube 2. The dispersed magnetic particles can make the biological reaction process more complete. When measuring the signal, the electromagnet 3 63 can cooperate with the electromagnet 1 61 and the electromagnet 2 62 to gather or disperse the magnetic particles, so as to improve the detection sensitivity and accuracy.

[0038] The optical signal detection device may be a photomultiplier tube or a high-sensitivity camera. The detection device is a prior art and will not be described in detail herein. The optical signal detection device is disposed above the capillary glass tube (not shown in the figure).

[0039] The process of using the device to perform chemiluminescent immunoassay is as follows:

[0040] 1) 10 μL of magnetic particle solution coated with antigen or antibody is added to the sample addition hole 11. The magnetic particle solution flows along the capillary glass tube 2 toward the hydrophobic ring 3 under capillary action. When it flows to the electromagnet 6, the magnetic particles will gather there, and the remaining solution will continue to flow forward to the hydrophobic ring 3 at the end of the capillary glass tube 2. Due to the hydrophobic effect, the liquid stops flowing and forms a convex liquid film based on surface tension. By setting the speed of the motor 5, the hydrophobic disk 4 eccentrically arranged relative to the hydrophobic ring 3 is rotated. The hydrophobic disk 4 destroys the surface tension of the liquid. The molecules on the surface of the hydrophobic disk drive the liquid to rotate, and the solution is gradually discharged from the end of the capillary glass tube 2 at a certain rate, so that the liquid can circulate in the tube;

[0041] 2) Add 10 μL of sample solution to the sample well 11. The sample solution flows along the capillary glass tube 2 toward the hydrophobic ring 3 under the action of capillaries. When it flows to the electromagnet 6, it will react with the antigen or antibody coated on the magnetic particles gathered there. During the reaction, the electromagnet 1 61 and the electromagnet 2 62 interact with each other under the drive of the alternating current, so that the magnetic particles gathered there are dispersed from each other, thereby enhancing the reaction efficiency. The solution after the reaction will continue to flow to the hydrophobic ring 3 at the end of the capillary glass tube 2. Set the speed of the motor 5 and rotate the hydrophobic disk 4 to discharge the solution in the tube at a certain rate.

[0042] 3) Add 50 μL of washing solution to the sample well 11. The washing solution flows along the capillary glass tube 2 toward the hydrophobic ring 3 under the action of capillaries, and flows to the electromagnet 6 to wash the magnetic particles gathered there. During the washing process, the electromagnet 1 61 and the electromagnet 2 62 interact with each other under the drive of the alternating current, so that the magnetic particles gathered there are dispersed from each other, thereby enhancing the washing efficiency. The washed solution continues to flow to the hydrophobic ring 3 at the end of the capillary glass tube 2. Set the speed of the motor 5, and rotate the hydrophobic disk 4 to discharge the solution in the tube at a certain rate.

[0043] 4) Add 10 μL of enzyme-labeled antibody solution to the sample well 11. The enzyme-labeled antibody solution flows along the capillary glass tube 2 toward the hydrophobic ring 3 under capillary action. When it flows to the electromagnet 6, it will react with the antigen or antibody coated on the magnetic particles gathered there. During the reaction, the electromagnet 1 61 and the electromagnet 2 62 interact with each other under the drive of the alternating current, so that the magnetic particles gathered there are dispersed from each other, thereby enhancing the reaction efficiency. The solution after the reaction will continue to flow to the hydrophobic ring 3 at the end of the capillary glass tube 2. Set the speed of the motor 5 and rotate the hydrophobic disk 4 to discharge the solution in the tube at a certain rate.

[0044] 5) Add 50 μL of washing solution to the sample well 11. The washing solution flows along the capillary glass tube 2 toward the hydrophobic ring 3 under the action of capillaries, and flows to the electromagnet 6 to wash the magnetic particles gathered there. During the washing process, the electromagnet 1 61 and the electromagnet 2 62 interact with each other under the drive of the alternating current, so that the magnetic particles gathered there are dispersed from each other, thereby enhancing the washing efficiency. The washed solution continues to flow to the hydrophobic ring 3 at the end of the capillary glass tube 2. Set the speed of the motor 5, and rotate the hydrophobic disk 4 to discharge the solution in the tube at a certain rate.

[0045] 6) Add 10 μL of enzymatic luminescent substrate solution to the sample well 11. The enzymatic luminescent substrate solution flows along the capillary glass tube 2 toward the hydrophobic ring 3 under the action of capillaries. When it flows to the electromagnet 6, it reacts with the enzyme bound to the magnetic particles gathered there. During the reaction, the electromagnet 1 61 and the electromagnet 2 62 interact with each other under the drive of the alternating current, so that the magnetic particles gathered there are dispersed from each other, thereby enhancing the reaction efficiency. The solution after the reaction continues to flow to the hydrophobic ring 3 at the end of the capillary glass tube 2. Set the speed of the motor 5 and rotate the hydrophobic disk 4 to discharge the solution in the tube at a certain rate.

[0046] 7) Signal detection: A photomultiplier tube or a high-sensitivity camera is provided at the electromagnet 6 and above the capillary glass tube 2 to collect light signals. At this time, the electromagnet three 63 cooperates with the electromagnet one 61 and the electromagnet two 62 to gather or disperse the magnetic particles, and analyzes the amount of the substance to be measured in the sample solution according to the intensity of the light signal. Example 2

[0047] The difference between this embodiment and embodiment 1 is that a capillary glass tube 2 is connected to two opposite sides of the sample adding block 1 to form two sets of detection paths. Of course, three sets of detection paths can also be designed based on the same sample adding block 1. When three sets of paths are designed, two adjacent capillary glass tubes 2 are arranged vertically. Based on this structure, two or three sets of parallel detection can be performed on the same sample at the same time, which helps to improve the accuracy and reliability of the detection results. The top view of the reaction device with two sets of detection paths is shown in FIG. Figure 7 shown.

[0048] The above shows and describes the basic principles, main features and advantages of the present invention. However, the above is only a specific embodiment of the present invention, and the technical features of the present invention are not limited thereto. Any other implementation methods derived by any technician in the field without departing from the technical solution of the present invention should be included in the patent scope of the present invention.

Claims

1. A microfluidic magnetic particle chemiluminescence reaction device, characterized in that: It includes a sample adding block, a capillary glass tube, a hydrophobic ring, a hydrophobic disk, a rotating driving mechanism and an electromagnet; The sample adding block is provided with a sample adding hole for dripping the reaction liquid; The capillary glass tube uses capillary action to guide the reaction liquid, and one end of the capillary glass tube is sealed and connected to the sample adding block and is connected to the sample adding hole; A hydrophobic ring is used to enhance the surface tension of the reaction liquid, and the hydrophobic ring is arranged on the outer peripheral surface of the end of the capillary glass tube away from the sample addition block; A hydrophobic disk, used to break the surface tension of the reaction liquid, the hydrophobic disk being connected to the rotary drive mechanism; An electromagnet is used to disperse and position the magnetic particles, and the electromagnet is arranged outside the capillary glass tube; The hydrophobic disk is arranged in parallel with the hydrophobic ring, and the distance between the hydrophobic ring and the hydrophobic disk is 0.01 to 0.50 mm; The hydrophobic disk is eccentrically arranged relative to the hydrophobic ring, and both the hydrophobic ring and the hydrophobic disk are made of hydrophobic material.

2. The microfluidic magnetic particle chemiluminescent reaction device according to claim 1, characterized in that: There are three electromagnets arranged outside the capillary glass tube, namely electromagnet one, electromagnet two and electromagnet three.

3. The microfluidic magnetic particle chemiluminescent reaction device according to claim 2, characterized in that: Electromagnet 1 and electromagnet 2 are respectively arranged at the front and rear sides of the capillary glass tube and are coaxially symmetrically arranged. Electromagnet 3 is arranged directly below the capillary glass tube, and the axis of electromagnet 3 is perpendicular to the axis of electromagnet 1.

4. The microfluidic magnetic particle chemiluminescent reaction device according to claim 1, characterized in that: The optical signal detection device is arranged above the capillary glass tube and corresponds to the position of the electromagnet.

5. The microfluidic magnetic particle chemiluminescent reaction device according to claim 1, characterized in that: The capillary glass tube is made of light-transmitting hydrophilic material.

6. The microfluidic magnetic particle chemiluminescent reaction device according to claim 1, characterized in that: The rotary drive mechanism is a motor, and the side of the hydrophobic disc away from the hydrophobic ring is coaxially fixedly connected to the motor shaft of the motor.

Images