Microfluidic device, fluid control method using same, and detection method using fluid control method

Abstract

The invention relates to a microfluidic device, a fluid control method using the microfluidic device, and a detection method using the fluid control method. The microfluidic device comprises a chip body and an air pressure control module, and the chip body comprises a flow channel layer, an air control layer, a diaphragm layer and the air pressure control module; the device is characterized in that the air control chamber comprises a split-flow check valve group, the air pressure control module is matched with the split-flow check valve group to independently control the on-off states of the reaction outflow control flow channel, the detection control flow channel and the liquid discharge control flow channel, and the split-flow check valve group has three states. When the shunting check valve group is in the second state, the liquid only flows to the waste liquid chamber from the reaction outflow control flow channel, so that the waste liquid is prevented from accidentally flowing into the detection cavity to influence the detection result; the liquid can be prevented from flowing back to the reaction chamber during flow division of the reaction flow-out control flow channel, and the device has the advantage of multiple purposes.

Description

Technical Field

[0001] This invention relates to the field of microfluidics, and more particularly to a pneumatic microfluidic device. Background Technology

[0002] Currently, the use of microfluidic ELISA technology to detect protein concentration is quite common. Immunoassay of protein requires 3-10 parallel runs for a single protein concentration. However, most commercially available microfluidic chips are single-channel or dual-channel, which require repeated sample loading, resulting in high time costs.

[0003] To address the aforementioned issues, existing technology includes a Chinese invention patent, ZL202410045114.1 (publication number CN117548164B), entitled "Microfluidic Chip, Detection Device and Method for Detecting the Properties and Flavors of Traditional Chinese Medicine," which discloses a microfluidic chip capable of simultaneously conducting four parallel experiments. This chip has four independently arranged microchannels. The sample is introduced into the capture chamber through the sample application chamber. The capture chamber, containing magnetic beads, is simultaneously connected to the detection chamber and the waste liquid chamber. The chip is mounted on a rotating device, which... The direction of rotation is used to control the flow of liquid from the capture chamber into the detection chamber or the waste liquid chamber. However, due to the large inertia of the detection chamber and the waste liquid chamber after high-speed rotation, when the rotation stops and the waste liquid is discharged or during detection, the liquid in the waste liquid chamber or the detection chamber interacts with the chamber wall and flows back into the flow channel, causing some magnetic beads to be undetectable and increasing the risk of false negatives. If the rotation speed is gradually reduced to avoid backflow, the time required for the deceleration process will be much longer than the time for the liquid to flow from the capture chamber into the next chamber, which will increase the number of operation steps and the experimental time.

[0004] Therefore, it is urgent to provide a microfluidic device that can prevent fluid from flowing back from the detection chamber to the reaction chamber and to separate the detection liquid and waste liquid, a fluid control method for use with the microfluidic device, and a detection method for detecting protein concentration using the fluid control method. Summary of the Invention

[0005] The first technical problem to be solved by the present invention is to provide a microfluidic device that prevents backflow and can separate waste liquid and detection liquid, in view of the above-mentioned existing technology.

[0006] The second technical problem to be solved by the present invention is to provide a fluid control method using the above-mentioned microfluidic device, in view of the current state of the prior art.

[0007] The third technical problem to be solved by the present invention is to provide a detection method that applies the above-mentioned fluid control method in view of the current state of the prior art.

[0008] The technical solution adopted by this invention to solve the first technical problem is as follows: the microfluidic device includes a chip body and a pressure control module, wherein the chip body includes:

[0009] The flow channel layer is constructed with a detection channel, which includes a reaction chamber, a detection chamber located downstream of and connected to the reaction chamber, and a waste liquid chamber located downstream of and connected to the reaction chamber.

[0010] The pneumatic control layer is constructed with a pneumatic control chamber, which covers a portion of the adjacent detection channel.

[0011] A diaphragm layer, which is a flexible thin film, is sealed between the gas control layer and the flow channel layer. When the gas control chamber is under negative pressure, the diaphragm layer deforms toward the gas control chamber, thereby forming a passage in the detection channel; when the gas control chamber is under positive pressure, the diaphragm layer deforms toward the detection channel, thereby forming an open circuit in the detection channel.

[0012] The air pressure control module is connected to the air control chamber and is used to control the air control chamber to be in a positive pressure state or a negative pressure state.

[0013] The feature is that: the reaction chamber extends downstream to form a reaction outflow control channel, and the downstream end of the reaction outflow control channel branches and extends into the detection chamber and the waste liquid chamber respectively, forming a detection control channel and a discharge control channel;

[0014] The pneumatic control chamber includes a diversion check valve assembly. The pneumatic pressure control module, in conjunction with the diversion check valve assembly, can independently control the on / off state of the reaction outflow control channel, the detection control channel, and the discharge control channel. The diversion check valve assembly has three states:

[0015] In the first state, the reaction outflow control channel is open / closed, the detection control channel is open, and the drain control channel is closed. Fluid flows out of the reaction chamber and through the reaction outflow control channel to the detection control channel.

[0016] In the second state, the reaction outflow control channel is open / closed, the detection control channel is closed, and the drain control channel is open. Fluid flows out of the reaction chamber and through the reaction outflow control channel to the drain control channel.

[0017] When in the closed state, the reaction outflow control channel is open / closed, the detection control channel is closed, and the drain control channel is closed.

[0018] To enhance the fluid transport capacity of the diversion check valve assembly, the diversion check valve assembly further includes:

[0019] The reaction outflow micro-valve chamber covers a portion of the reaction outflow control channel and is used to control the on / off state of the reaction outflow control channel;

[0020] The liquid storage micropump chamber covers a portion of the reaction outflow control channel and is used to control the on / off state of the reaction outflow control channel. The coverage area of ​​the liquid storage micropump chamber over the reaction outflow control channel is greater than the coverage area of ​​the reaction outflow microvalve chamber over the reaction outflow control channel.

[0021] The flow into the micro-valve chamber is detected, which corresponds to a portion of the detection and control flow channel, and is used to control the on / off state of the detection and control flow channel;

[0022] Waste liquid flows into the micro-valve chamber, correspondingly covering a portion of the drain control channel, and is used to control the on / off state of the drain control channel. The pneumatic control module independently controls the pneumatic pressure of the storage micro-pump chamber, the detection inflow micro-valve chamber, and the waste liquid inflow micro-valve chamber. The storage micro-pump chamber increases the pressure difference between the reaction outflow control channel and the detection control channel or drain control channel, increasing the kinetic energy of the fluid movement, thereby allowing the liquid to fully flow into the detection control channel or drain control channel.

[0023] To simplify the flow channel structure of the detection channel, the number of reaction chambers in the detection channel is two, and the number of waste liquid chambers is one; the drainage control channel converges from the two reaction chambers to the waste liquid chamber into a single "T" shape.

[0024] Furthermore, the diversion check valve assembly includes two interconnected micropump chambers for liquid storage, two interconnected microvalve chambers for reaction outflow, two microvalve chambers for detecting inflow, and one microvalve chamber for waste liquid inflow. The waste liquid inflow microvalve chamber corresponds to the coverage of the drain control channel and is located downstream of the convergence point of the drain control channel. This technical solution reduces the number of waste liquid chambers distributed on the channel layer.

[0025] To prevent backflow of liquid in the reaction chamber, the flow channel layer is further constructed with a sample inlet channel, which is connected to the detection channel by the reaction inflow channel.

[0026] Furthermore, the gas control chamber also includes a check valve assembly, which corresponds to the reaction inflow channel and can work with the gas pressure control module to control the on / off state of the reaction inflow channel. The ratio of the number of check valve assemblies to the number of diversion check valve assemblies in the detection channel is: [ratio to be inserted here]. Users can prevent liquid in the reaction chamber from flowing back into the sample inlet channel by closing the check valve assembly, thus reducing the occurrence of false negatives.

[0027] To further increase the fluid transport capacity of the check valve assembly, the check valve assembly further includes:

[0028] The liquid inlet micro-valve chamber covers a portion of the reaction inflow channel;

[0029] The liquid inlet micropump chamber is located downstream of the liquid inlet microvalve chamber, correspondingly covering a portion of the reaction outflow control channel, and the coverage area of ​​the liquid inlet micropump chamber over the reaction inflow channel is greater than the coverage area of ​​the liquid inlet micropump chamber over the reaction inflow channel.

[0030] The reaction inflow microvalve chamber is located downstream of the liquid inlet micropump chamber, correspondingly covering a portion of the reaction outflow control channel. The gas pressure control module independently controls the gas pressure of the liquid inlet microvalve chamber, the liquid inlet micropump chamber, and the reaction inflow microvalve chamber. The liquid inlet micropump chamber creates a pressure difference at each covered point within the reaction inflow channel, enabling independent pressure differentials to drive fluid into the reaction chamber. Furthermore, when closed, it prevents fluid backflow into the sample inlet channel.

[0031] To enable users to control the flow of fluid within the injection chamber, the injection channel further includes an injection chamber for introducing reaction liquid, sample, and washing liquid. The gas control layer also includes a sample loading gas control chamber connected to a gas pressure control module. The gas pressure control module can control the sample loading gas control chamber to be in a positive or negative pressure state, and the sample loading gas control chamber covers a portion of the injection channel.

[0032] Furthermore, the sample injection chamber includes a first liquid storage chamber, a second liquid storage chamber, a third liquid storage chamber, and a fourth liquid storage chamber. The first, second, third, and fourth liquid storage chambers respectively extend downstream to form a first, second, third, and fourth sample dispensing channel, respectively. Correspondingly, the sample dispensing gas control chamber includes:

[0033] The first sample dispensing micro-valve chamber covers a portion of the first sample dispensing channel and is used to control the on / off state of the first sample dispensing channel;

[0034] The second sample dispensing micro-valve chamber covers a portion of the second sample dispensing channel and is used to control the on / off state of the second sample dispensing channel;

[0035] The third sample dispensing micro-valve chamber covers a portion of the third sample dispensing channel and is used to control the on / off state of the third sample dispensing channel;

[0036] The fourth sample dispensing micro-valve chamber covers a portion of the fourth sample dispensing channel and is used to control the on / off state of the fourth sample dispensing channel.

[0037] To enable simultaneous testing of multiple sets of experiments, the flow channel layer is further constructed with multiple detection channels arranged along the fluid movement direction of the detection channels. Correspondingly, each of the following is provided with multiple inlet microvalve chambers, inlet micropump chambers, reaction inflow microvalve chambers, reaction outflow microvalve chambers, storage micropump chambers, detection inflow microvalve chambers, and waste liquid inflow microvalve chambers:

[0038] Each of the liquid inlet micro-valve chambers is connected from left to right along the direction of fluid movement perpendicular to the detection channel to form a first air path. The first air path also has a first air inlet connected to the air pressure control module.

[0039] Each of the liquid inlet micropump chambers is connected from left to right along the direction of fluid movement perpendicular to the detection channel to form a second air passage. The second air passage also has a second air inlet connected to the air pressure control module.

[0040] A third gas path is formed between each of the reaction inflow microvalve chambers, which is connected from left to right along the direction of fluid movement perpendicular to the detection channel. The third gas path also has a third air inlet connected to the gas pressure control module.

[0041] A fourth gas path is formed by connecting each of the reaction outflow microvalve chambers from left to right along the direction of fluid movement perpendicular to the detection channel. The fourth gas path also has a fourth air inlet connected to the gas pressure control module.

[0042] Each of the liquid storage micropump chambers is connected from left to right along the direction of fluid movement perpendicular to the detection channel to form a fifth air path, and the fifth air path also has a fifth air inlet connected to the air pressure control module;

[0043] A sixth air passage is formed by connecting each of the detection inflow microvalve chambers from left to right along the direction of fluid movement perpendicular to the detection channel. The sixth air passage also has a sixth air inlet connected to the air pressure control module.

[0044] A seventh air path is formed between the various waste liquid inflow micro-valve chambers, which are connected from left to right along the fluid movement direction perpendicular to the detection channel. The seventh air path also has a seventh air inlet connected to the air pressure control module.

[0045] In addition, the gas control layer also forms a first sample dispensing gas path that is connected to the first sample dispensing micro-valve chamber. The first sample dispensing gas path has a first sample dispensing inlet that is connected to the gas pressure control module on the surface of the gas control layer.

[0046] The gas control layer also forms a second sample dispensing gas path that is connected to the second sample dispensing microvalve chamber. The second sample dispensing gas path has a second sample dispensing inlet that is connected to the gas pressure control module on the surface of the gas control layer.

[0047] The gas control layer also forms a third sample dispensing gas path that is connected to the third sample dispensing micro-valve chamber. The third sample dispensing gas path has a third sample dispensing inlet that is connected to the gas pressure control module on the surface of the gas control layer.

[0048] The gas control layer also forms a fourth sample dispensing gas path that communicates with the fourth sample dispensing microvalve chamber. This fourth sample dispensing gas path has a fourth sample inlet on its surface that communicates with the gas pressure control module. Changing the positive or negative gas pressure in a single gas path allows for simultaneous control of the microvalve chambers or micropump chambers located on that gas path for different detection channels, simplifying the user's operation.

[0049] The technical solution adopted by the present invention to solve the second technical problem is as follows: the fluid control method is characterized by: the application of the above-mentioned microfluidic device, and includes the following steps:

[0050] a) Initialization: The sample injection chamber contains fluid, the gas pressure control module controls each gas path of the gas control layer to be in a positive pressure state, the check valve group and the diversion check valve group are both in a closed state, and the sample injection channel and the detection channel are both in an open circuit state.

[0051] b) Sample addition to the reaction chamber: Open the check valve assembly, the sample inlet channel is connected to the reaction inlet channel, and the fluid from step a flows from the sample inlet chamber to the reaction chamber. After the fluid has completely flowed into the reaction chamber, close the check valve assembly.

[0052] c) Waste liquid discharge from the reaction chamber: The pneumatic control module switches the diversion check valve group from the closed state to the second state. The fluid in step b flows out of the reaction chamber and flows through the reaction outflow control channel to the discharge control channel. After the fluid has completely flowed into the waste liquid chamber, the diversion check valve group is switched to the closed state.

[0053] d) Inflow into the detection chamber: The pneumatic control module switches the diversion check valve assembly from the closed state to the first state. The fluid from step b flows out of the reaction chamber, through the reaction outflow control channel, and into the detection control channel. After the fluid has completely flowed into the detection chamber, the diversion check valve assembly is switched back to the closed state. By switching the states of the check valve assembly and the diversion check valve assembly, the flow direction of the liquid in the reaction chamber is controlled. The actions of closing the check valve assembly and switching the diversion check valve assembly to the closed state can prevent the backflow of fluid in the reaction chamber, detection chamber, and waste liquid chamber.

[0054] Users can provide a discharge port on the surface of the flow channel layer that communicates with the detection chamber to discharge the fluid inside the detection chamber, or they can use a diversion check valve assembly to push the fluid inside the detection chamber to the waste liquid chamber. Preferably, the fluid control method further includes the following steps:

[0055] f) Waste liquid discharge from the detection chamber: Control the diversion check valve group to the first state. At this time, the reaction outflow control channel is open / closed, the detection control channel is open, and the discharge control channel is closed. Then switch the second gas path from the positive pressure state to the negative pressure state. The fluid in step d flows from the detection chamber to the reaction outflow control channel.

[0056] After all the fluid has flowed into the reaction outflow control channel, the diversion check valve assembly is switched from the first state to the second state. The fluid then flows from the reaction outflow control channel to the waste liquid chamber. Once the fluid has completely flowed into the waste liquid chamber, the diversion check valve assembly is switched to the closed state. The user pushes the fluid in the detection chamber to the waste liquid chamber by switching the state of the diversion check valve assembly. A pressure difference is generated within the detection channel to drive the fluid movement, eliminating the need for additional fluid input to drive the discharge. This method offers the advantages of precise control and low reagent consumption.

[0057] In order to ensure that the liquid in the injection channel is evenly drawn into each reaction chamber, the operation of opening the check valve assembly further includes the following steps:

[0058] g1) Switch the first gas path from a positive pressure state to a negative pressure state. At this time, the first gas path is in a negative pressure state, the second gas path is in a positive pressure state, and the third gas path is in a positive pressure state. The fluid begins to move towards the reaction inflow channel.

[0059] g2) Switch the second gas path from the positive pressure state in step g1 to the negative pressure state. At this time, the first gas path is in the negative pressure state, the second gas path is in the negative pressure state, and the third gas path is in the positive pressure state. The fluid flows into the reaction flow channel.

[0060] g3) Switch the first gas path from the negative pressure state of step g2 to the positive pressure state, and switch the third gas path from the positive pressure state of step g to the negative pressure state. At this time, the first gas path is in the positive pressure state, the second gas path is in the negative pressure state, and the third gas path is in the negative pressure state, and the fluid begins to flow into the reaction chamber.

[0061] g4) Switch the second gas path from the negative pressure state in step g3 to the positive pressure state. At this time, the first gas path is in the positive pressure state, the second gas path is in the positive pressure state, and the third gas path is in the negative pressure state. The fluid accelerates its flow to the reaction chamber.

[0062] The operation of closing the check valve assembly specifically includes the following steps:

[0063] g5) The pressure control module controls the first, second, and third gas paths to be in a positive pressure state. The state switching of the second gas path creates a pressure difference between the corresponding coverage point of the reaction inflow channel and other coverage points within the reaction inflow channel, thereby giving the fluid strong kinetic energy as it flows from the injection channel to the reaction chamber.

[0064] To ensure that the fluid in the reaction chamber can flow fully into the detection chamber or waste chamber, the operation of switching the diversion check valve assembly from the closed state to the first state further includes the following steps:

[0065] h1) Switch the fourth gas path from a positive pressure state to a negative pressure state. At this time, the fourth gas path is in a negative pressure state, the fifth gas path is in a positive pressure state, the sixth gas path is in a positive pressure state, and the seventh gas path is in a positive pressure state. The fluid starts to flow from the reaction chamber to the reaction outlet control channel.

[0066] h2) Switch the fifth gas path from a positive pressure state to a negative pressure state. At this time, the fourth gas path is in a negative pressure state, the fifth gas path is in a negative pressure state, the sixth gas path is in a positive pressure state, and the seventh gas path is in a positive pressure state. The fluid in step h1 flows into the reaction outflow control channel.

[0067] h3) Switch the fourth gas path from negative pressure to positive pressure and the sixth gas path from positive pressure to negative pressure. At this time, the fourth gas path is in positive pressure, the fifth gas path is in negative pressure, the sixth gas path is in negative pressure, and the seventh gas path is in positive pressure. The fluid in step h2 begins to flow from the reaction outlet control channel to the detection control channel.

[0068] h4) Switch the fifth gas path from a negative pressure state to a positive pressure state. At this time, the fourth gas path is in a positive pressure state, the fifth gas path is in a positive pressure state, the sixth gas path is in a negative pressure state, and the seventh gas path is in a positive pressure state. The fluid in step h3 flows into the detection control channel and flows into the detection chamber.

[0069] The operation of switching the diversion check valve assembly from the closed state to the second state includes the following steps:

[0070] i1) Switch the fourth gas path from a positive pressure state to a negative pressure state. At this time, the fourth gas path is in a negative pressure state, the fifth gas path is in a positive pressure state, the sixth gas path is in a positive pressure state, and the seventh gas path is in a positive pressure state. The fluid starts to flow from the reaction chamber to the reaction outlet control channel.

[0071] i2) Switch the fifth gas path from a positive pressure state to a negative pressure state. At this time, the fourth gas path is in a negative pressure state, the fifth gas path is in a negative pressure state, the sixth gas path is in a positive pressure state, and the seventh gas path is in a positive pressure state. The fluid in step i1 flows into the reaction outflow control channel.

[0072] i3) Switch the fourth gas path from negative pressure to positive pressure and the seventh gas path from positive pressure to negative pressure. At this time, the fourth gas path is in positive pressure, the fifth gas path is in negative pressure, the sixth gas path is in positive pressure, and the seventh gas path is in negative pressure. The fluid in step i2 begins to flow from the reaction outlet control channel to the discharge control channel.

[0073] i4) The fifth gas path is switched from a negative pressure state to a positive pressure state. At this time, the fourth gas path is in a positive pressure state, the fifth gas path is in a positive pressure state, the sixth gas path is in a positive pressure state, and the seventh gas path is in a negative pressure state. The fluid from step i3 flows into the detection and control channel and then flows to the waste liquid chamber. The state switching of the fifth gas path creates a pressure difference between the reaction inflow channel and the corresponding coverage point of the fifth gas path and the reaction chamber, detection chamber, and waste liquid chamber, thereby giving the fluid strong kinetic energy when flowing from the reaction chamber to the detection chamber or waste liquid chamber.

[0074] To ensure that the fluid in the detection chamber can flow fully into the waste liquid chamber, the control of the diversion check valve assembly to be in the first state further includes the following steps:

[0075] j) Switch the sixth gas path from a positive pressure state to a negative pressure state. At this time, the fourth gas path is in a positive pressure state, the fifth gas path is in a positive pressure state, the sixth gas path is in a negative pressure state, and the seventh gas path is in a positive pressure state. The fluid starts to flow from the detection chamber to the detection control channel.

[0076] The operation of switching the diversion check valve assembly from the first state to the second state includes the following steps:

[0077] k1) Switch the fifth gas path from a positive pressure state to a negative pressure state. At this time, the fourth gas path is in a positive pressure state, the fifth gas path is in a negative pressure state, the sixth gas path is in a negative pressure state, and the seventh gas path is in a positive pressure state. The fluid in step j flows from the detection control channel into the reaction outflow control channel.

[0078] k2) After the fluid from step j has completely flowed into the reaction outflow control channel, the sixth gas path is switched from a negative pressure state to a positive pressure state, and the seventh gas path is switched from a positive pressure state to a negative pressure state. At this time, the fourth gas path is in a positive pressure state, the fifth gas path is in a negative pressure state, the sixth gas path is in a positive pressure state, and the seventh gas path is in a negative pressure state. The fluid from step k1 begins to flow from the reaction outflow control channel into the detection control channel.

[0079] (k3) Switch the fifth gas path from a negative pressure state to a positive pressure state. At this time, the fourth gas path, the fifth gas path, the sixth gas path are in a positive pressure state, and the seventh gas path is in a negative pressure state. The fluid from step k2 flows from the reaction outlet control channel into the waste liquid chamber through the detection control channel. This technical solution cleverly utilizes the state switching of the fifth gas path to create a pressure difference between the reaction outlet control channel and the detection chamber, causing the fluid in the detection chamber to flow into the reaction outlet control channel. During this period, the fourth gas path is in a positive pressure state to ensure that the fluid in the detection chamber does not enter the reaction chamber. A pressure difference is created between the reaction outlet control channel and the waste liquid chamber, causing the fluid to flow from the reaction outlet control channel to the waste liquid chamber. During this period, the sixth gas path is in a positive pressure state to ensure that the fluid does not flow back into the detection chamber.

[0080] The technical solution adopted by the present invention to solve the third technical problem is as follows: The detection method is characterized by the application of the above-mentioned fluid control method, wherein the reaction chamber of the microfluidic device contains magnetic beads, and the microfluidic device further includes a magnetic adsorption module corresponding to the reaction chamber. The magnetic adsorption module can turn the magnetic field on or off to achieve the adsorption of the magnetic beads on the bottom wall of the reaction chamber or suspension in the fluid of the reaction chamber. The surface of the magnetic beads has an anti-acid, and the method includes the following steps:

[0081] S1 Sample Injection Chamber Filling: Perform the initialization operation as described in step a on the microfluidic device, wherein the first reservoir contains the protein to be tested and the enzyme-labeled secondary antibody solution, the second reservoir contains the colorimetric solution, and the third reservoir contains the stop solution.

[0082] Formation of S2 magnetic bead-antibody-test protein conjugate: The pressure control module controls the first sample dispensing gas path to be in a negative pressure state, and then performs the sample dispensing operation in the reaction chamber in step b. The test protein and enzyme-labeled secondary antibody solution in step S1 flow into the reaction chamber from the first storage chamber and are incubated with the magnetic beads in the reaction chamber to obtain a magnetic bead-antibody-test protein conjugate mixture.

[0083] S3 Magnetic Separation Waste Discharge: The magnetic adsorption module is activated, and the magnetic bead-antibody-test protein conjugate mixture obtained in step S2 is magnetically separated into magnetic bead-antibody-test protein conjugate adsorbed to the bottom wall of the reaction chamber and the first waste liquid; the reaction chamber waste discharge operation in step c is performed, and the first waste liquid flows from the reaction chamber into the waste liquid chamber;

[0084] S4 Color Development: The pressure control module controls the second sample loading gas path to be in a negative pressure state before performing the reaction chamber sample loading operation of operation b. The color developing solution flows into the reaction chamber from the second storage chamber. After the color developing solution has completely flowed into the reaction chamber, the second sample loading gas path is controlled to be in a positive pressure state, and the magnetic adsorption module is turned off. The color developing solution reacts with the magnetic bead-antibody-test protein conjugate obtained in step S3 in the reaction chamber to obtain the color developing product.

[0085] S5 Termination: The pressure control module controls the third sample addition gas path to be in a negative pressure state, and then performs the sample addition operation of the reaction chamber in operation b. The termination liquid flows into the reaction chamber from the third storage chamber. After the termination liquid has completely flowed into the reaction chamber, the third sample addition gas path is controlled to be in a positive pressure state. The termination liquid reacts with the colorimetric product obtained in step S4 in the reaction chamber to obtain the product to be detected.

[0086] S6 Detection and Analysis: Following step d, the product to be detected flows into the detection chamber. The product obtained in step S5 flows from the reaction chamber into the detection chamber. The microfluidic device also includes a detection module for detecting optical signals within the detection chamber. The detection module is activated to detect the product in the detection chamber and obtain absorbance values. The absorbance values ​​obtained from each detection channel are then averaged to obtain the average absorbance value, which is converted to a protein concentration value using a standard curve method. This technical solution utilizes a gas path to control the liquid path flow of the flow channel layer, enabling simultaneous acquisition of detection results from each detection channel and improving experimental efficiency.

[0087] To facilitate subsequent experiments within the chip itself, the fourth liquid storage chamber further stores washing liquid, and the following steps are included after step S6:

[0088] S7 Washing: The pressure control module controls the fourth sample dispensing gas path to be under negative pressure before performing the sample dispensing operation in the reaction chamber (step b). The washing liquid flows into the reaction chamber from the fourth storage chamber. After the stop liquid has completely flowed into the reaction chamber, the fourth sample dispensing gas path is controlled to be under positive pressure, and the flow into the detection chamber (step d) is executed. The washing liquid flows from the reaction chamber into the detection chamber. The waste liquid discharge operation in the detection chamber (step f) is then executed, and the mixed waste liquid of the product to be tested and the washing liquid obtained in step S6 flows from the detection chamber into the waste liquid chamber. By executing the waste liquid discharge operation in the detection chamber (step f), the user can simultaneously clean the detection chamber and the waste liquid chamber at the bifurcation end in one washing step, simplifying the washing operation.

[0089] Compared with the prior art, the advantages of the present invention are as follows:

[0090] 1. The pneumatic control module can independently control the flow path and disconnection status of the reaction outflow control channel, the detection control channel, and the discharge control channel. When the diversion check valve group is closed, the flow between the reaction chamber, the detection chamber, and the waste liquid chamber is interrupted, preventing backflow into the channel between the three chambers. When the diversion check valve group is in the first state, the liquid will only flow from the reaction outflow control channel to the detection chamber, preventing the liquid from accidentally flowing into the waste liquid chamber after the reaction and reducing the risk of false negative test results.

[0091] 2. When the diversion check valve assembly is in the second state, the liquid will only flow from the reaction outflow control channel to the waste liquid chamber, avoiding the waste liquid from accidentally flowing into the detection chamber and affecting the detection results. In addition, the reaction outflow control channel is open and closed in the first and second states, which can prevent the liquid in the reaction outflow control channel from flowing back to the reaction chamber detection liquid diversion when diverting, and has the advantage of "one thing for multiple uses".

[0092] 3. Users can push the fluid in the detection chamber to the waste liquid chamber by switching the state of the diversion check valve group. The pressure difference generated inside the detection channel drives the fluid movement. No other fluid needs to be input to drive the discharge. It has the advantages of precise control and low reagent consumption.

[0093] 4. The detection method of the present invention uses the gas path to control the liquid path of the flow channel layer, which can simultaneously obtain the detection results of each detection channel and improve the experimental efficiency. Attached Figure Description

[0094] Figure 1 This is a schematic diagram of the chip body structure according to Embodiment 1 of the present invention;

[0095] Figure 2 This is an exploded view of the chip body structure according to Embodiment 1 of the present invention;

[0096] Figure 3 This is a schematic diagram showing the positional relationship between the flow channel layer and the gas control layer at the coverage area in Embodiment 1 of the present invention;

[0097] Figure 4 This is a schematic diagram of the flow channel layer in Embodiment 1 of the present invention;

[0098] Figure 5 This is a schematic diagram of the gas control layer in Embodiment 1 of the present invention;

[0099] Figure 6 This is a control relationship diagram between the air pressure control module and the air control room in Embodiment 1 of the present invention. Detailed Implementation

[0100] The present invention will be further described in detail below with reference to the accompanying drawings and embodiments.

[0101] Example 1

[0102] like Figures 1-6 The diagram shows a preferred embodiment of the present invention. The microfluidic device of this embodiment is mainly used for protein detection. The detection process requires sample injection, washing, drainage, and delivery of the final product into the detection chamber for detection. During this process, because all flow channels are identical, liquid within the chip often flows back or cross-flows into other chambers, causing the detection process to fail. Therefore, providing a microfluidic device that prevents backflow and can separate waste liquid and detection liquid would be beneficial. The following will provide a detailed description of this microfluidic device:

[0103] See Figures 1 to 5The microfluidic device includes a chip body 1 and a gas pressure control module. The chip body 1 includes a flow channel layer 2, a gas control layer 3, and a diaphragm layer 20. The flow channel layer 2 is configured with a detection channel 21, which includes a reaction chamber 211, a detection chamber 22 located downstream of and connected to the reaction chamber 211, and a waste liquid chamber 23 located downstream of and connected to the reaction chamber 211. The gas control layer 3 is configured with a gas control chamber 30, which covers a portion of the adjacent detection channel 21. The diaphragm layer 20 is sealed. A flexible thin film, diaphragm 20, is positioned between the pneumatic control layer 3 and the flow channel layer 2. When the pneumatic control chamber 30 is under negative pressure, the diaphragm 20 deforms towards the inside of the pneumatic control chamber 30, thus forming a passage within the detection channel 21. When the pneumatic control chamber 30 is under positive pressure, the diaphragm 20 deforms towards the inside of the detection channel 21, thus forming a break within the detection channel 21. The pneumatic pressure control module is connected to the pneumatic control chamber 30 and is used to control whether the pneumatic control chamber 30 is under positive or negative pressure. A reaction outflow control channel 212 extends downstream from the reaction chamber 211. Downstream of 212, a detection control channel 213 and a discharge control channel 214 extend and branch out towards the detection chamber 22 and waste liquid chamber 23, respectively. The pneumatic control chamber 30 includes a diversion check valve assembly 31. The pneumatic control module, in conjunction with the diversion check valve assembly 31, can independently control the on / off states of the reaction outflow control channel 212, the detection control channel 213, and the discharge control channel 214. The diversion check valve assembly 31 has three states: in the first state, the reaction outflow control channel 212 is open / closed, the detection control channel 213 is open, and the discharge channel 214 is closed. When the control channel 214 is disconnected, fluid flows out of the reaction chamber 211 and through the reaction outflow control channel 212 to the detection control channel 213. In the second state, the reaction outflow control channel 212 is open / closed, the detection control channel 213 is disconnected, and the drain control channel 214 is open, allowing fluid to flow out of the reaction chamber 211 and through the reaction outflow control channel 212 to the drain control channel 214. When the system is closed, the reaction outflow control channel 212 is open / closed, the detection control channel 213 is disconnected, and the drain control channel 214 is disconnected. Additionally, multiple screw holes are provided on the chip body 1, each containing a screw. Users can tighten these screws to prevent leakage of liquid and gas from the flow channel layer 2 and the gas control layer 3.

[0104] Please refer to Figure 6The pneumatic control module includes a solenoid valve assembly 11 for controlling the pneumatic pressure state of the pneumatic control chamber 30, a compressor pump 12 for providing negative pressure, a vacuum pump 13 for providing positive pressure, and a controller 14 connected to the solenoid valve assembly for controlling the opening and closing state of the solenoid valve assembly. The solenoid valve assembly has a first pneumatic pressure input port, a second pneumatic pressure input port, and a pneumatic pressure output port, wherein the pneumatic pressure output port is connected to each air inlet of the pneumatic control chamber 30, the first pneumatic pressure input port is connected to the compressor pump 12, and the second pneumatic pressure input port is connected to the vacuum pump 13. The solenoid valve assembly 11 includes a first solenoid valve 1101 connected to the first sample inlet 331 and a second sample inlet 34. The following are connected to the following: a second solenoid valve 1102 connected to the first air inlet 41; a third solenoid valve 1103 connected to the third sample inlet 351; a fourth solenoid valve 1104 connected to the fourth sample inlet 361; a fifth solenoid valve 1105 connected to the first air inlet 41; a sixth solenoid valve 1106 connected to the second air inlet 51; a seventh solenoid valve 1107 connected to the third air inlet 61; an eighth solenoid valve 1108 connected to the fourth air inlet 71; a ninth solenoid valve 1109 connected to the fifth air inlet 81; a tenth solenoid valve 1110 connected to the sixth air inlet 91; and an eleventh solenoid valve 1111 connected to the seventh air inlet 101.

[0105] The diversion check valve assembly 31 includes a reaction outflow microvalve chamber 311, a liquid storage micropump chamber 312, an inflow detection microvalve chamber 313, and a waste liquid inflow microvalve chamber 314. The reaction outflow microvalve chamber 311 covers a portion of the reaction outflow control channel 212 and is used to control the on / off state of the reaction outflow control channel 212. The liquid storage micropump chamber 312 also covers a portion of the reaction outflow control channel 212 and is used to control the on / off state of the reaction outflow control channel 212. The liquid storage micropump chamber 312 controls the on / off state of the reaction outflow control channel 212. The coverage area is larger than the coverage area of ​​the reaction outflow microvalve chamber 311 over the reaction outflow control channel 212; the detection inflow microvalve chamber 313 covers a portion of the detection control channel 213 and is used to control the on / off state of the detection control channel 213; the waste liquid inflow microvalve chamber 314 covers a portion of the drain control channel 214 and is used to control the on / off state of the drain control channel 214; the pneumatic pressure control module independently controls the pneumatic pressure of the storage micropump chamber 312, the detection inflow microvalve chamber 313, and the waste liquid inflow microvalve chamber 314. The setting of the storage micropump chamber 312 increases the pressure difference between the reaction outflow control channel 212 and the detection control channel 213 or the drain control channel 214, increases the kinetic energy of the fluid movement, and thus allows the liquid to fully flow into the detection control channel 213 or the drain control channel 214, enhancing the fluid transport capacity of the diversion check valve assembly 31.

[0106] The detection channel 21 has two reaction chambers 211 and one waste liquid chamber 23. The drain control channel 214 converges from the two reaction chambers 211 to the waste liquid chamber 23 in a "T" shape. Correspondingly, the diversion check valve group 31 includes two interconnected storage micro pump chambers 312, two interconnected reaction outflow micro valve chambers 311, two detection inflow micro valve chambers 313, and one waste liquid inflow micro valve chamber 314. The waste liquid inflow micro valve chamber 314 is located downstream of the convergence point of the drain control channel 214, corresponding to the coverage of the drain control channel 214. This technical solution reduces the number of waste liquid chambers 23 distributed on the channel layer 2 and simplifies the channel structure of the detection channel 21.

[0107] The flow channel layer 2 also includes a sample inlet channel 24 for sample introduction. The sample inlet channel 24 is connected to the detection channel 21 by a reaction inflow channel 25. The pneumatic control chamber 30 also includes a check valve assembly 32, which corresponds to the reaction inflow channel 25 and can work with the pneumatic pressure control module to control the on / off state of the reaction inflow channel 25. The ratio of the number of check valve assemblies 32 to the number of diversion check valve assemblies 31 in the detection channel 21 is 2:1. By closing the check valve assembly 32, the user can prevent the liquid in the reaction chamber 211 from flowing back into the sample inlet channel 24, thereby reducing the occurrence of false negatives and preventing backflow of liquid in the reaction chamber 211.

[0108] The check valve assembly 32 includes an inlet micro-valve chamber 321, an inlet micro-pump chamber 322, and a reaction inflow micro-valve chamber 323. The inlet micro-valve chamber 321 covers a portion of the reaction inflow channel 25. The inlet micro-pump chamber 322 is located downstream of the inlet micro-valve chamber 321 and covers a portion of the reaction outflow control channel 212. The coverage area of ​​the inlet micro-pump chamber 322 over the reaction inflow channel 25 is greater than the coverage area of ​​the reaction inflow channel 25. The reaction inflow micro-valve chamber 323 is located downstream of the inlet micro-pump chamber 322 and covers a portion of the reaction outflow control channel 212. The pressure control module independently controls the pressure of the inlet micro-valve chamber 321, the inlet micro-pump chamber 322, and the reaction inflow micro-valve chamber 323. The micropump chamber 322 creates a pressure difference at each covered point in the reaction inflow channel 25, which can generate an independent pressure difference in the reaction inflow channel 25 to push the fluid into the reaction chamber 211. When closed, it can also prevent the fluid from flowing back into the sample inlet channel 24, thus increasing the fluid transport capacity of the check valve assembly 32.

[0109] The sample inlet channel 24 includes a sample inlet chamber 240 for introducing reaction solution, sample and washing solution. The gas control layer 3 is also constructed with a sample loading gas control chamber 300 connected to the gas pressure control module. The gas pressure control module can control the sample loading gas control chamber 300 to be in a positive pressure state or a negative pressure state. The sample loading gas control chamber 300 covers a part of the sample inlet channel 24, which allows the user to control the flow of fluid in the sample inlet chamber 240.

[0110] The sample injection chamber 240 includes a first liquid storage chamber 241, a second liquid storage chamber 242, a third liquid storage chamber 243, and a fourth liquid storage chamber 244. The first liquid storage chamber 241, the second liquid storage chamber 242, the third liquid storage chamber 243, and the fourth liquid storage chamber 244 extend downstream to form corresponding first sample dispensing channels 245, second sample dispensing channels 246, third sample dispensing channels 247, and fourth sample dispensing channels 248, respectively. Correspondingly, the sample dispensing pneumatic control chamber 300 includes a first sample dispensing microvalve chamber 301, a second sample dispensing microvalve chamber 302, a third sample dispensing microvalve chamber 303, and a fourth sample dispensing microvalve chamber 304. The first sample dispensing microvalve chamber 301 covers a portion of the first sample dispensing channel 245 and is used to control the on / off state of the first sample dispensing channel 245; the second sample dispensing microvalve chamber 302 covers a portion of the second sample dispensing channel 246 and is used to control the on / off state of the second sample dispensing channel 246; the third sample dispensing microvalve chamber 303 covers a portion of the third sample dispensing channel 247 and is used to control the on / off state of the third sample dispensing channel 247; and the fourth sample dispensing microvalve chamber 304 covers a portion of the fourth sample dispensing channel 248 and is used to control the on / off state of the fourth sample dispensing channel 248. Additionally, the upper surface of the channel layer 2 has a first sample inlet communicating with the first liquid storage chamber 241, a second sample inlet communicating with the second liquid storage chamber 242, a third sample inlet communicating with the third liquid storage chamber 243, and a fourth sample inlet communicating with the fourth liquid storage chamber 244. The upper surface of the channel layer 2 also has a waste liquid outlet communicating with the waste liquid chamber 23.

[0111] Finally, the flow channel layer is constructed with multiple detection channels 21 arranged along the fluid movement direction of the detection channel 21. Correspondingly, multiple inlet micro-valve chambers 321, inlet micro-pump chambers 322, reaction inflow micro-valve chambers 323, reaction outflow micro-valve chambers 311, storage micro-pump chambers 312, detection inflow micro-valve chambers 313, and waste liquid inflow micro-valve chambers 314 are each provided.

[0112] Each liquid inlet micro-valve chamber 321 is connected from left to right along the fluid movement direction perpendicular to the detection channel 21 to form a first air passage 4. The first air passage 4 also has a first air inlet 41 connected to the air pressure control module.

[0113] Each liquid inlet micropump chamber 322 is connected from left to right along the fluid movement direction perpendicular to the detection channel 21 to form a second air passage 5. The second air passage 5 also has a second air inlet 51 connected to the air pressure control module.

[0114] Each reaction flows into the micro-valve chamber 323 and is connected from left to right along the fluid movement direction perpendicular to the detection channel 21 to form a third gas path 6. The third gas path 6 also has a third air inlet 61 connected to the gas pressure control module.

[0115] Each reaction outlet microvalve chamber 311 is connected from left to right along the fluid movement direction perpendicular to the detection channel 21 to form a fourth gas path 7. The fourth gas path 7 also has a fourth air inlet 71 connected to the gas pressure control module.

[0116] Each liquid storage micropump chamber 312 is connected from left to right along the direction of fluid movement perpendicular to the detection channel 21 to form a fifth air passage 8. The fifth air passage 8 also has a fifth air inlet 81 connected to the air pressure control module.

[0117] A sixth air passage 9 is formed by connecting each detection inlet microvalve chamber 313 from left to right along the direction of fluid movement perpendicular to the detection channel 21. The sixth air passage 9 also has a sixth air inlet 91 connected to the pressure control module. It is observed that the sixth air passage 9 also covers a part of the drain control channel 214. This may cause the drain control channel 214 to open or close when the sixth air passage 9 is in a negative pressure state or a positive pressure state, so that the fluid in the reaction chamber 211 cannot flow in a directional manner. To address this problem, this embodiment can provide a rigid interlocking block between the channel layer 2 and the diaphragm layer 20 at the covering part to counteract the force of deformation of the diaphragm layer 20 caused by the change of air pressure in the sixth air passage 9.

[0118] The waste liquid flows into the micro valve chamber 314 and connects from left to right along the fluid movement direction perpendicular to the detection channel 21 to form a seventh air path 10. The seventh air path 10 also has a seventh air inlet 101 connected to the air pressure control module.

[0119] In addition, the gas control layer 3 also forms a first sample dispensing gas path 33 that is connected to the first sample dispensing micro valve chamber 301. The first sample dispensing gas path 33 forms a first sample dispensing inlet 331 on the surface of the gas control layer 3 that is connected to the gas pressure control module.

[0120] The gas control layer 3 also forms a second sample dispensing gas path 34 that is connected to the second sample dispensing micro valve chamber 302. The second sample dispensing gas path 34 has a second sample dispensing inlet 341 that is connected to the gas pressure control module on the surface of the gas control layer 3.

[0121] The gas control layer 3 also constitutes a third sample delivery gas path 35 that is connected to the third sample delivery micro valve chamber 303. The third sample delivery gas path 35 has a third sample delivery inlet 351 that is connected to the gas pressure control module on the surface of the gas control layer 3.

[0122] The gas control layer 3 also forms a fourth sample dispensing gas path 36, which is connected to the fourth sample dispensing microvalve chamber 304. The fourth sample dispensing gas path 36 has a fourth sample dispensing inlet 361 on the surface of the gas control layer 3, which is connected to the gas pressure control module. By changing the positive or negative gas pressure of a single gas path, the microvalve chambers or micropump chambers located on the gas path of different detection channels 21 can be controlled simultaneously, simplifying the user's operation steps and enabling the user to perform multiple sets of detection experiments at the same time.

[0123] The working principle of the microfluidic device in this embodiment is as follows: the user adds different solutions to the first liquid storage chamber 241, the second liquid storage chamber 242, the third liquid storage chamber 243 and the fourth liquid storage chamber 244; the check valve group 32 is opened, and the sample injection chamber 240 adds a sample to the reaction chamber 211; the diversion check valve group 31 is switched to the first state, and the liquid to be tested in the reaction chamber 211 flows into the detection chamber 22; the diversion check valve 31 is switched to the second state, and the waste liquid in the reaction chamber 211 flows into the waste liquid chamber 23, thereby realizing the diversion of waste liquid and detection liquid.

[0124] Example 2: A fluid control method

[0125] The fluid control method of this embodiment uses the microfluidic device of Embodiment 1 and includes the following steps:

[0126] a) Initialization: The sample injection chamber 240 contains fluid. The gas pressure control module controls each gas path of the gas control layer 3 to be in a positive pressure state. The check valve group 32 and the diversion check valve group 31 are both in the closed state. The sample injection channel 24 and the detection channel 21 are both in the open circuit state.

[0127] b) Adding sample to the reaction chamber: Open the check valve assembly 32, and the sample inlet channel 24 and the reaction inlet channel 25 are connected. The fluid from step a flows from the sample inlet chamber 240 to the reaction chamber 211. After the fluid has completely flowed into the reaction chamber 211, close the check valve assembly 32.

[0128] c) Waste liquid discharge from reaction chamber: The pneumatic control module switches the diversion check valve group 31 from the closed state to the second state. The fluid in step b flows out of the reaction chamber 211 and flows through the reaction outflow control channel 212 to the discharge control channel 214. After the fluid has completely flowed into the waste liquid chamber 23, the diversion check valve group 31 is switched to the closed state.

[0129] d) Inflow into the detection chamber: The pneumatic control module switches the diversion check valve assembly 31 from the closed state to the first state. The fluid from step b flows out of the reaction chamber 211, through the reaction outflow control channel 212, and into the detection control channel 213. After the fluid has completely flowed into the detection chamber 22, the diversion check valve assembly 31 is switched back to the closed state. By switching the states of the check valve assembly 32 and the diversion check valve assembly 31, the flow direction of the liquid in the reaction chamber 211 is controlled. The actions of closing the check valve assembly 32 and switching the diversion check valve assembly 31 to the closed state can prevent the backflow of fluid in the reaction chamber 211, the detection chamber 22, and the waste liquid chamber 23.

[0130] Users can provide a discharge port on the surface of the flow channel layer 2 that communicates with the detection chamber 22 to discharge the fluid in the detection chamber 22, or they can use the diversion check valve assembly 31 to push the fluid in the detection chamber 22 to the waste liquid chamber 23. Preferably, the fluid control method further includes the following steps:

[0131] f) Waste liquid discharge from the detection chamber: The control diversion check valve group 31 is in the first state. At this time, the reaction outflow control channel 212 is open and closed, the detection control channel 213 is open, and the discharge control channel 214 is closed. Then, the second gas path 5 is switched from the positive pressure state to the negative pressure state. The fluid in step d flows from the detection chamber 22 to the reaction outflow control channel 212.

[0132] After all the fluid has flowed into the reaction outflow control channel 212, the diversion check valve assembly 31 is switched from the first state to the second state. The fluid flows from the reaction outflow control channel 212 to the waste liquid chamber 23. After the fluid has completely flowed into the waste liquid chamber 23, the diversion check valve assembly 31 is switched to the closed state. The user pushes the fluid in the detection chamber 22 to the waste liquid chamber 23 by switching the state of the diversion check valve assembly 31. A pressure difference is generated inside the detection channel 21 to drive the fluid movement. No other fluid needs to be input to drive the discharge, which has the advantages of precise control and low reagent consumption.

[0133] The operation of opening the check valve assembly 32 specifically includes the following steps:

[0134] g1) Switch the first gas path 4 from a positive pressure state to a negative pressure state. At this time, the first gas path 4 is in a negative pressure state, the second gas path 5 is in a positive pressure state, the third gas path 6 is in a positive pressure state, and the fluid begins to move towards the reaction inflow channel 25.

[0135] g2) Switch the second gas path 5 from the positive pressure state in step g1 to the negative pressure state. At this time, the first gas path 4 is in the negative pressure state, the second gas path 5 is in the negative pressure state, and the third gas path 6 is in the positive pressure state. The fluid flows into the reaction inflow channel 25.

[0136] g3) Switch the first gas path 4 from the negative pressure state of step g2 to the positive pressure state, and switch the third gas path 6 from the positive pressure state of step g2 to the negative pressure state. At this time, the first gas path 4 is in the positive pressure state, the second gas path 5 is in the negative pressure state, and the third gas path 6 is in the negative pressure state, and the fluid begins to flow into the reaction chamber 211.

[0137] g4) Switch the second gas path 5 from the negative pressure state in step g3 to the positive pressure state. At this time, the first gas path 4 is in the positive pressure state, the second gas path 5 is in the positive pressure state, and the third gas path 6 is in the negative pressure state. The fluid accelerates and flows towards the reaction chamber 211.

[0138] The operation of closing the check valve assembly 32 specifically includes the following steps:

[0139] (g5) The pressure control module controls the first gas path 4, the second gas path 5, and the third gas path 6 to be in a positive pressure state. The state switching of the second gas path 5 creates a pressure difference between the corresponding covered area of ​​the reaction inflow channel 25 and other covered areas within the reaction inflow channel 25, thereby giving the fluid strong kinetic energy as it flows from the injection channel 24 to the reaction chamber 211, thus enabling the liquid in the injection channel 24 to be uniformly drawn into each reaction chamber 211.

[0140] The operation of switching the diversion check valve assembly 31 from the closed state to the first state includes the following steps:

[0141] h1) Switch the fourth gas path 7 from a positive pressure state to a negative pressure state. At this time, the fourth gas path 7 is in a negative pressure state, the fifth gas path 8 is in a positive pressure state, the sixth gas path 9 is in a positive pressure state, and the seventh gas path 10 is in a positive pressure state. The fluid flows from the reaction chamber 211 to the reaction outflow control channel 212.

[0142] h2) Switch the fifth gas path 8 from a positive pressure state to a negative pressure state. At this time, the fourth gas path 7 is in a negative pressure state, the fifth gas path 8 is in a negative pressure state, the sixth gas path 9 is in a positive pressure state, and the seventh gas path 10 is in a positive pressure state. The fluid in step h1 flows into the reaction outflow control channel 212.

[0143] h3) Switch the fourth gas path 7 from negative pressure to positive pressure and the sixth gas path 9 from positive pressure to negative pressure. At this time, the fourth gas path 7 is in positive pressure, the fifth gas path 8 is in negative pressure, the sixth gas path 9 is in negative pressure, and the seventh gas path 10 is in positive pressure. The fluid in step h2 begins to flow from the reaction outlet control channel 212 to the detection control channel 213.

[0144] h4) Switch the fifth gas path 8 from negative pressure to positive pressure. At this time, the fourth gas path 7 is in positive pressure, the fifth gas path 8 is in positive pressure, the sixth gas path 9 is in negative pressure, and the seventh gas path 10 is in positive pressure. The fluid in step h3 flows into the detection control channel 213 and flows to the detection chamber 22.

[0145] The operation of switching the diversion check valve assembly 31 from the closed state to the second state includes the following steps:

[0146] i1) Switch the fourth gas path 7 from a positive pressure state to a negative pressure state. At this time, the fourth gas path 7 is in a negative pressure state, the fifth gas path 8 is in a positive pressure state, the sixth gas path 9 is in a positive pressure state, and the seventh gas path 10 is in a positive pressure state. The fluid flows from the reaction chamber 211 to the reaction outlet control channel 212.

[0147] i2) Switch the fifth gas path 8 from a positive pressure state to a negative pressure state. At this time, the fourth gas path 7 is in a negative pressure state, the fifth gas path 8 is in a negative pressure state, the sixth gas path 9 is in a positive pressure state, and the seventh gas path 10 is in a positive pressure state. The fluid in step i1 flows into the reaction outflow control channel 212.

[0148] i3) Switch the fourth gas path 7 from negative pressure to positive pressure and the seventh gas path 10 from positive pressure to negative pressure. At this time, the fourth gas path 7 is in positive pressure, the fifth gas path 8 is in negative pressure, the sixth gas path 9 is in positive pressure, and the seventh gas path 10 is in negative pressure. The fluid in step i2 begins to flow from the reaction outlet control channel 212 to the liquid discharge control channel 214.

[0149] i4) Switch the fifth gas path 8 from a negative pressure state to a positive pressure state. At this time, the fourth gas path 7, the fifth gas path 8, and the sixth gas path 9 are in a positive pressure state, and the seventh gas path 10 is in a negative pressure state. The fluid from step i3 flows into the detection and control flow channel 213 and then into the waste liquid chamber 23. The state switching of the fifth gas path 8 creates a pressure difference between the corresponding coverage area of ​​the reaction inflow channel 25 and the fifth gas path 8 and the reaction chamber 211, the detection chamber 22, and the waste liquid chamber 23. This results in the fluid having strong kinetic energy as it flows from the reaction chamber 211 to the detection chamber 22 or the waste liquid chamber 23, allowing the fluid in the reaction chamber 211 to fully flow into the detection chamber 22 or the waste liquid chamber 23.

[0150] Controlling the flow diversion check valve assembly 31 to be in the first state includes the following steps:

[0151] j) Switch the sixth gas path 7 from a positive pressure state to a negative pressure state. At this time, the fourth gas path 7 is in a positive pressure state, the fifth gas path 8 is in a positive pressure state, the sixth gas path 9 is in a negative pressure state, and the seventh gas path 10 is in a positive pressure state. The fluid starts to flow from the detection chamber 22 to the detection control channel 213.

[0152] The operation of switching the diversion check valve assembly 31 from the first state to the second state includes the following steps:

[0153] k1) Switch the fifth gas path 8 from a positive pressure state to a negative pressure state. At this time, the fourth gas path 7 is in a positive pressure state, the fifth gas path 8 is in a negative pressure state, the sixth gas path 9 is in a negative pressure state, and the seventh gas path 10 is in a positive pressure state. The fluid in step j flows from the detection control channel 213 into the reaction outflow control channel 212.

[0154] k2) After the fluid from step j has completely flowed into the reaction outflow control channel 212, the sixth gas path 9 is switched from a negative pressure state to a positive pressure state, and the seventh gas path 10 is switched from a positive pressure state to a negative pressure state. At this time, the fourth gas path 7 is in a positive pressure state, the fifth gas path 8 is in a negative pressure state, the sixth gas path 9 is in a positive pressure state, and the seventh gas path 10 is in a negative pressure state. The fluid from step k1 begins to flow from the reaction outflow control channel 212 into the detection control channel 213.

[0155] (k3) Switch the fifth gas path 8 from a negative pressure state to a positive pressure state. At this time, the fourth gas path 7, the fifth gas path 8, and the sixth gas path 9 are in a positive pressure state, and the seventh gas path 10 is in a negative pressure state. The fluid from step k2 flows from the reaction outflow control channel 212 into the waste liquid chamber 23 via the detection control channel 213. This technical solution cleverly utilizes the state switching of the fifth gas path 8 to create a pressure difference between the reaction outflow control channel 212 and the detection chamber 22, causing the fluid in the detection chamber 22 to flow into the reaction outflow control channel 212. During this period, the fourth gas path 7 is in a positive pressure state to ensure that the fluid in the detection chamber 22 does not enter the reaction chamber 211. The pressure difference created between the reaction outflow control channel 212 and the waste liquid chamber 22 causes the fluid to flow from the reaction outflow control channel 212 to the waste liquid chamber 23. During this period, the sixth gas path 9 is in a positive pressure state to ensure that the fluid does not flow back into the detection chamber 22, thereby allowing the fluid in the detection chamber 22 to fully flow into the waste liquid chamber 23.

[0156] Example 3: A detection method

[0157] The detection method of this embodiment applies the fluid control method of Embodiment 2. The reaction chamber 211 of the microfluidic device contains magnetic beads. The microfluidic device also includes a magnetic adsorption module corresponding to the reaction chamber 211. The magnetic adsorption module can turn the magnetic field on or off to enable the magnetic beads to be adsorbed onto the bottom wall of the reaction chamber 211 or suspended in the fluid of the reaction chamber 211. The surface of the magnetic beads has an anti-acid. The method includes the following steps:

[0158] S1 Sample Injection Chamber Filling: Perform the initialization operation of the microfluidic device as in step a, wherein the first reservoir 241 stores the protein to be tested and the enzyme-labeled secondary antibody solution, the second reservoir 242 stores the colorimetric solution, and the third reservoir 243 stores the stop solution.

[0159] Formation of S2 magnetic bead-antibody-test protein conjugate: The pressure control module controls the first sample dispensing gas path 33 to be in a negative pressure state, and then the reaction chamber sample dispensing operation in step b is performed. The test protein and enzyme-labeled secondary antibody solution in step S1 flow from the first storage chamber 241 into the reaction chamber 211 and are incubated with the magnetic beads in the reaction chamber 211 to obtain the magnetic bead-antibody-test protein conjugate mixture.

[0160] S3 Magnetic Separation Waste Discharge: Activate the magnetic adsorption module. The magnetic bead-antibody-test protein conjugate mixture obtained in step S2 is magnetically separated into magnetic bead-antibody-test protein conjugate adsorbed onto the bottom wall of reaction chamber 211 and the first waste liquid. Perform the reaction chamber waste discharge operation in step c. The first waste liquid flows from reaction chamber 211 into waste liquid chamber 23.

[0161] S4 Color Development: The pressure control module controls the second sample loading gas path 34 to be in a negative pressure state, and then the reaction chamber sample loading operation of operation b is performed. The color development solution flows into the reaction chamber 211 from the second storage chamber 242. After the color development solution has completely flowed into the reaction chamber 211, the second sample loading gas path 34 is controlled to be in a positive pressure state, the magnetic adsorption module is closed, and the color development solution reacts with the magnetic bead-antibody-test protein conjugate obtained in step S3 in the reaction chamber 211 to obtain the color development product.

[0162] S5 Termination: The pressure control module controls the third sample addition gas path 35 to be in a negative pressure state, and then performs the sample addition operation of the reaction chamber in operation b. The stop liquid flows into the reaction chamber 211 from the third liquid storage chamber 243. After the stop liquid has completely flowed into the reaction chamber 211, the third sample addition gas path 35 is controlled to be in a positive pressure state. The stop liquid and the colorimetric product obtained in step S4 react in the reaction chamber 211 to obtain the product to be detected.

[0163] S6 Detection and Analysis: Following step d, the product to be detected flows into the detection chamber. The product obtained in step S5 flows from reaction chamber 211 into detection chamber 22. The microfluidic device also includes a detection module for detecting optical signals within the detection chamber. The detection module is activated to detect the product in detection chamber 22 and obtain absorbance values. The absorbance values ​​obtained from each detection channel 21 are then averaged to obtain the average absorbance value, which is converted to a protein concentration value using a standard curve method. This technical solution utilizes a gas path to control the flow path of the liquid path in the flow channel layer 2, enabling simultaneous acquisition of detection results from each detection channel 21, thus improving experimental efficiency.

[0164] The fourth storage chamber 244 contains washing liquid, and the following steps are included after step S6:

[0165] S7 Washing: The gas pressure control module controls the fourth sample dispensing gas path 36 to be in a negative pressure state before performing the reaction chamber sample dispensing operation (step b). The washing liquid flows from the fourth storage chamber 244 into the reaction chamber 211. After the stop liquid has completely flowed into the reaction chamber 211, the fourth sample dispensing gas path 36 is controlled to be in a positive pressure state, and the flow into the detection chamber operation (step d) is executed. The washing liquid flows from the reaction chamber 211 into the detection chamber 22. The detection chamber waste liquid discharge operation (step f) is executed, and the mixed waste liquid of the product to be detected and the washing liquid obtained in step S6 flows from the detection chamber 22 into the waste liquid chamber 23. By executing the detection chamber waste liquid discharge operation (step f), the user can simultaneously clean the detection chamber 22 and the waste liquid chamber 23 at the bifurcation end in one washing step, simplifying the washing operation steps. After washing, it is convenient for the user to perform the next experiment in the chip body 1.

Claims

1. A microfluidic device, comprising a chip body (1) and a pressure control module, wherein the chip body (1) comprises: The flow channel layer (2) is constructed with a detection channel (21), the detection channel (21) including a reaction chamber (211), a detection chamber (22) located downstream of the reaction chamber (211) and connected to the reaction chamber (211), and a waste liquid chamber (23) located downstream of the reaction chamber (211) and connected to it. The reaction chamber (211) is located inside the reaction chamber (211). The gas control layer (3) is constructed with a gas control chamber (30), which covers a portion of the adjacent detection channel (21); A diaphragm layer (20) is sealed between the pneumatic control layer (3) and the flow channel layer (2) and is a flexible thin film. When the pneumatic control chamber (30) is under negative pressure, the diaphragm layer (20) deforms toward the pneumatic control chamber (30) to form a passage in the detection channel (21); when the pneumatic control chamber (30) is under positive pressure, the diaphragm layer (20) deforms toward the detection channel (21) to form an open circuit in the detection channel (21). The air pressure control module is connected to the air control chamber (30) and is used to control the air control chamber (30) to be in a positive pressure state or a negative pressure state. Its features are: The reaction chamber (211) extends downstream to form a reaction outflow control channel (212), and the downstream end of the reaction outflow control channel (212) branches and extends into the detection chamber (22) and the waste liquid chamber (23) respectively, forming a detection control channel (213) and a discharge control channel (214). The pneumatic control chamber (30) includes a diversion check valve assembly (31). The pneumatic pressure control module, in conjunction with the diversion check valve assembly (31), can independently control the on / off state of the reaction outflow control channel (212), the detection control channel (213), and the discharge control channel (214). The diversion check valve assembly (31) has three states: In the first state, the reaction outflow control channel (212) is open and closed, the detection control channel (213) is open, and the drain control channel (214) is closed. The fluid flows out of the reaction chamber (211) and flows through the reaction outflow control channel (212) to the detection control channel (213). In the second state, the reaction outflow control channel (212) is open and closed, the detection control channel (213) is closed, and the drain control channel (214) is open. The fluid flows out of the reaction chamber (211) and flows through the reaction outflow control channel (212) to the drain control channel (214). When in the closed state, the reaction outflow control channel (212) is open, the detection control channel (213) is closed, and the drain control channel (214) is closed.

2. The microfluidic device according to claim 1, characterized in that... The aforementioned diversion check valve assembly (31) includes: The reaction outflow micro-valve chamber (311) covers a portion of the reaction outflow control channel (212) and is used to control the on / off state of the reaction outflow control channel (212); The liquid storage micropump chamber (312) covers a portion of the reaction outflow control channel (212) and is used to control the on / off state of the reaction outflow control channel (212). The coverage area of ​​the liquid storage micropump chamber (312) over the reaction outflow control channel (212) is greater than the coverage area of ​​the reaction outflow microvalve chamber (311) over the reaction outflow control channel (212). The flow into the micro-valve chamber (313) is detected, which corresponds to a portion of the detection control channel (213), and is used to control the on / off state of the detection control channel (213); Waste liquid flows into the micro-valve chamber (314), correspondingly covering a portion of the drain control channel (214), and is used to control the on / off state of the drain control channel (214); The pressure control module independently controls the pressure of the liquid storage micropump chamber (312), the detection flow microvalve chamber (313), and the waste liquid flow microvalve chamber (314).

3. The microfluidic device according to claim 2, characterized in that: The number of reaction chambers (211) on the detection channel (21) is two, and the number of waste liquid chambers (23) is one; the drain control channel (214) converges from the two reaction chambers (211) to the waste liquid chamber (23) into a "T" shape.

4. The microfluidic device according to claim 3, characterized in that: The diversion check valve assembly (31) includes two interconnected storage micropump chambers (312), two interconnected reaction outflow microvalve chambers (311), two detection inflow microvalve chambers (313), and a waste liquid inflow microvalve chamber (314). The waste liquid inflow microvalve chamber (314) corresponds to the coverage of the drain control channel (214) and is located downstream of the confluence of the drain control channel (214).

5. The microfluidic device according to claim 4, characterized in that: The flow channel layer (2) is also constructed with a sample inlet channel (24) for sample introduction, and the sample inlet channel (24) is connected to the detection channel (21) by a reaction inlet flow channel (25).

6. The microfluidic device according to claim 5, characterized in that: The pneumatic control chamber (30) also includes a check valve assembly (32), which corresponds to the reaction inflow channel (25) and can cooperate with the pneumatic pressure control module to control the on / off state of the reaction inflow channel (25). The ratio of the number of check valve assemblies (32) to the number of diversion check valve assemblies (31) in the detection channel (21) is 2:

1.

7. The microfluidic device according to claim 6, characterized in that... The check valve assembly (32) includes: The liquid inlet micro-valve chamber (321) covers a portion of the reaction inlet channel (25); The liquid inlet micropump chamber (322) is located downstream of the liquid inlet microvalve chamber (321), correspondingly covering a part of the reaction outflow control channel (212), and the coverage area of ​​the liquid inlet micropump chamber (322) over the reaction inflow channel (25) is greater than the coverage area of ​​the liquid inlet micropump chamber (322) over the reaction inflow channel (25). The reaction flows into the microvalve chamber (323), which is located downstream of the liquid inlet micropump chamber (322) and covers a portion of the reaction outflow control channel (212). The gas pressure control module controls the gas pressure of the liquid inlet microvalve chamber (321), the liquid inlet micropump chamber (322) and the reaction flowing into the microvalve chamber (323) independently.

8. The microfluidic device according to claim 7, characterized in that: The sample inlet channel (24) includes an inlet chamber (240) for introducing reaction solution, sample and washing solution. The gas control layer (3) also has a sample loading gas control chamber (300) connected to the gas pressure control module. The gas pressure control module can control the sample loading gas control chamber (300) to be in a positive pressure state or a negative pressure state. The sample loading gas control chamber (300) covers a part of the sample inlet channel (24).

9. The microfluidic device according to claim 8, characterized in that: The sample injection chamber (240) includes a first liquid storage chamber (241), a second liquid storage chamber (242), a third liquid storage chamber (243), and a fourth liquid storage chamber (244). The first liquid storage chamber (241), the second liquid storage chamber (242), the third liquid storage chamber (243), and the fourth liquid storage chamber (244) extend downstream to form corresponding first sample dispensing channels (245), second sample dispensing channels (246), third sample dispensing channels (247), and fourth sample dispensing channels (248), respectively. Correspondingly, the sample dispensing gas control chamber (300) includes: The first sample dispensing micro-valve chamber (301) covers a portion of the first sample dispensing channel (245) and is used to control the on / off state of the first sample dispensing channel (245); The second sample dispensing micro-valve chamber (302) covers a portion of the second sample dispensing channel (246) and is used to control the on / off state of the second sample dispensing channel (246); The third sample dispensing micro-valve chamber (303) covers a portion of the third sample dispensing channel (247) and is used to control the on / off state of the third sample dispensing channel (247); The fourth sample dispensing micro-valve chamber (304) covers a portion of the fourth sample dispensing channel (248) and is used to control the on / off state of the fourth sample dispensing channel (248).

10. The microfluidic device according to claim 9, characterized in that: The flow channel layer is constructed with multiple detection channels (21) arranged along the fluid movement direction of the detection channel (21). Correspondingly, each of the following is provided with multiple inlet micro-valve chambers (321), inlet micro-pump chambers (322), reaction inflow micro-valve chambers (323), reaction outflow micro-valve chambers (311), storage micro-pump chambers (312), detection inflow micro-valve chambers (313), and waste liquid inflow micro-valve chambers (314). Each of the liquid inlet micro-valve chambers (321) is connected from left to right along the fluid movement direction perpendicular to the detection channel (21) to form a first air passage (4). The first air passage (4) also has a first air inlet (41) connected to the air pressure control module. Each of the liquid inlet micropump chambers (322) is connected from left to right along the fluid movement direction perpendicular to the detection channel (21) to form a second air passage (5). The second air passage (5) also has a second air inlet (51) connected to the air pressure control module. Each of the reaction inflow microvalve chambers (323) is connected from left to right along the fluid movement direction perpendicular to the detection channel (21) to form a third gas path (6), and the third gas path (6) also has a third air inlet (61) connected to the gas pressure control module; A fourth gas path (7) is formed between each of the reaction outflow microvalve chambers (311) from left to right along the fluid movement direction perpendicular to the detection channel (21). The fourth gas path (7) also has a fourth air inlet (71) connected to the air pressure control module. Each of the liquid storage micropump chambers (312) is connected from left to right along the fluid movement direction perpendicular to the detection channel (21) to form a fifth air passage (8). The fifth air passage (8) also has a fifth air inlet (81) connected to the air pressure control module. The detection flow microvalve chambers (313) are connected from left to right along the direction of fluid movement perpendicular to the detection channel (21) to form a sixth air passage (9). The sixth air passage (9) also has a sixth air inlet (91) connected to the air pressure control module. The waste liquid flows into the micro valve chamber (314) and connects from left to right along the fluid movement direction perpendicular to the detection channel (21) to form a seventh air path (10). The seventh air path (10) also has a seventh air inlet (101) connected to the air pressure control module. In addition, the gas control layer (3) also constitutes a first sample dispensing gas path (33) connected to the first sample dispensing micro valve chamber (301), and the first sample dispensing gas path (33) has a first sample dispensing inlet (331) connected to the gas pressure control module on the surface of the gas control layer (3); The gas control layer (3) also constitutes a second sample dispensing gas path (34) connected to the second sample dispensing micro valve chamber (302). The second sample dispensing gas path (34) has a second sample dispensing inlet (341) connected to the gas pressure control module on the surface of the gas control layer (3). The gas control layer (3) also constitutes a third sample dispensing gas path (35) connected to the third sample dispensing micro valve chamber (303). The third sample dispensing gas path (35) has a third sample dispensing inlet (351) connected to the gas pressure control module on the surface of the gas control layer (3). The gas control layer (3) also constitutes a fourth sample dispensing gas path (36) connected to the fourth sample dispensing micro valve chamber (304). The fourth sample dispensing gas path (36) has a fourth sample dispensing inlet (361) connected to the gas pressure control module on the surface of the gas control layer (3).

11. A fluid control method, characterized in that: The application of the microfluidic device as described in claim 10 includes the following steps: a) Initialization: The sample injection chamber (240) contains fluid, the gas pressure control module controls each gas path of the gas control layer (3) to be in a positive pressure state, the check valve group (32) and the diversion check valve group (31) are both in a closed state, and the sample injection channel (24) and the detection channel (21) are both in an open circuit state. b) Adding sample to the reaction chamber: Open the check valve assembly (32), the sample inlet channel (24) is connected to the reaction inlet channel (25), and the fluid in step a flows from the sample inlet chamber (240) to the reaction chamber (211). After the fluid has completely flowed into the reaction chamber (211), close the check valve assembly (32). c) Waste liquid discharge from the reaction chamber: The pneumatic control module switches the diversion check valve group (31) from the closed state to the second state. The fluid in step b flows out from the reaction chamber (211) and flows through the reaction outflow control channel (212) to the discharge control channel (214). After the fluid has completely flowed into the waste liquid chamber (23), the diversion check valve group (31) is switched to the closed state. d) Flow into the detection chamber: The pneumatic control module switches the diversion check valve group (31) from the closed state to the first state. The fluid in step b flows out of the reaction chamber (211) and flows through the reaction outflow control channel (212) to the detection control channel (213). After the fluid has completely flowed into the detection chamber (22), the diversion check valve group (31) is switched to the closed state.

12. The fluid control method according to claim 11, characterized in that, It also includes the following steps: f) Waste liquid discharge from the detection chamber: Control the diversion check valve group (31) to be in the first state. At this time, the reaction outflow control channel (212) is open and closed, the detection control channel (213) is open, and the discharge control channel (214) is closed. Then switch the second gas path (5) from the positive pressure state to the negative pressure state. The fluid in step d flows from the detection chamber (22) to the reaction outflow control channel (212). After all the fluids have flowed into the reaction outflow control channel (212), the diversion check valve group (31) is switched from the first state to the second state. The fluids flow from the reaction outflow control channel (212) to the waste liquid chamber (23). After all the fluids have flowed into the waste liquid chamber (23), the diversion check valve group (31) is switched to the closed state.

13. The fluid control method according to claim 12, characterized in that: The operation of opening the check valve assembly (32) specifically includes the following steps: g1) Switch the first gas path (4) from a positive pressure state to a negative pressure state. At this time, the first gas path (4) is in a negative pressure state, the second gas path (5) is in a positive pressure state, and the third gas path (6) is in a positive pressure state. The fluid begins to move towards the reaction inflow channel (25). g2) Switch the second gas path (5) from the positive pressure state in step g1 to the negative pressure state. At this time, the first gas path (4) is in the negative pressure state, the second gas path (5) is in the negative pressure state, and the third gas path (6) is in the positive pressure state. The fluid flows into the reaction inflow channel (25). g3) Switch the first gas path (4) from the negative pressure state of step g2 to the positive pressure state, and switch the third gas path (6) from the positive pressure state of step g2 to the negative pressure state. At this time, the first gas path (4) is in the positive pressure state, the second gas path (5) is in the negative pressure state, and the third gas path (6) is in the negative pressure state. The fluid begins to flow into the reaction chamber (211). g4) Switch the second gas path (5) from the negative pressure state in step g3 to the positive pressure state. At this time, the first gas path (4) is in the positive pressure state, the second gas path (5) is in the positive pressure state, and the third gas path (6) is in the negative pressure state. The fluid accelerates and flows towards the reaction chamber (211). The operation of closing the check valve assembly (32) specifically includes the following steps: The pressure control module described in g5) controls the first air path (4), the second air path (5), and the third air path (6) to be in a positive pressure state.

14. The fluid control method according to claim 13, characterized in that... The operation of switching the diversion check valve assembly (31) from the closed state to the first state includes the following steps: h1) Switch the fourth gas path (7) from a positive pressure state to a negative pressure state. At this time, the fourth gas path (7) is in a negative pressure state, the fifth gas path (8) is in a positive pressure state, the sixth gas path (9) is in a positive pressure state, and the seventh gas path (10) is in a positive pressure state. The fluid starts to flow from the reaction chamber (211) to the reaction outlet control channel (212). h2) Switch the fifth gas path (8) from a positive pressure state to a negative pressure state. At this time, the fourth gas path (7) is in a negative pressure state, the fifth gas path (8) is in a negative pressure state, the sixth gas path (9) is in a positive pressure state, and the seventh gas path (10) is in a positive pressure state. The fluid in step h1 flows into the reaction outflow control channel (212). h3) Switch the fourth gas path (7) from negative pressure to positive pressure, and switch the sixth gas path (9) from positive pressure to negative pressure. At this time, the fourth gas path (7) is in positive pressure, the fifth gas path (8) is in negative pressure, the sixth gas path (9) is in negative pressure, and the seventh gas path (10) is in positive pressure. The fluid in step h2 begins to flow from the reaction outlet control channel (212) to the detection control channel (213). h4) Switch the fifth gas path (8) from negative pressure to positive pressure. At this time, the fourth gas path (7) is in positive pressure, the fifth gas path (8) is in positive pressure, the sixth gas path (9) is in negative pressure, and the seventh gas path (10) is in positive pressure. The fluid in step h3 flows into the detection control channel (213) and flows to the detection chamber (22). The operation of switching the diversion check valve assembly (31) from the closed state to the second state includes the following steps: i1) Switch the fourth gas path (7) from a positive pressure state to a negative pressure state. At this time, the fourth gas path (7) is in a negative pressure state, the fifth gas path (8) is in a positive pressure state, the sixth gas path (9) is in a positive pressure state, and the seventh gas path (10) is in a positive pressure state. The fluid starts to flow from the reaction chamber (211) to the reaction outlet control channel (212). i2) Switch the fifth gas path (8) from a positive pressure state to a negative pressure state. At this time, the fourth gas path (7) is in a negative pressure state, the fifth gas path (8) is in a negative pressure state, the sixth gas path (9) is in a positive pressure state, and the seventh gas path (10) is in a positive pressure state. The fluid in step i1 flows into the reaction outflow control channel (212). i3) Switch the fourth gas path (7) from negative pressure to positive pressure, and switch the seventh gas path (10) from positive pressure to negative pressure. At this time, the fourth gas path (7) is in positive pressure, the fifth gas path (8) is in negative pressure, the sixth gas path (9) is in positive pressure, and the seventh gas path (10) is in negative pressure. The fluid in step i2 begins to flow from the reaction outlet control channel (212) to the liquid discharge control channel (214). i4) Switch the fifth gas path (8) from negative pressure to positive pressure. At this time, the fourth gas path (7) is in positive pressure, the fifth gas path (8) is in positive pressure, the sixth gas path (9) is in positive pressure, and the seventh gas path (10) is in negative pressure. The fluid in step i3 flows into the detection and control channel (213) and flows to the waste liquid chamber (23).

15. The fluid control method according to claim 14, characterized in that... The process of controlling the diversion check valve assembly (31) to be in the first state includes the following steps: j) Switch the sixth gas path (7) from a positive pressure state to a negative pressure state. At this time, the fourth gas path (7) is in a positive pressure state, the fifth gas path (8) is in a positive pressure state, the sixth gas path (9) is in a negative pressure state, and the seventh gas path (10) is in a positive pressure state. The fluid flows from the detection chamber (22) to the detection control channel (213). The operation of switching the diversion check valve assembly (31) from the first state to the second state includes the following steps: k1) Switch the fifth gas path (8) from a positive pressure state to a negative pressure state. At this time, the fourth gas path (7) is in a positive pressure state, the fifth gas path (8) is in a negative pressure state, the sixth gas path (9) is in a negative pressure state, and the seventh gas path (10) is in a positive pressure state. The fluid in step j flows from the detection control channel (213) into the reaction outflow control channel (212). k2) After the fluid from step j has completely flowed into the reaction outflow control channel (212), the sixth gas path (9) is switched from a negative pressure state to a positive pressure state, and the seventh gas path (10) is switched from a positive pressure state to a negative pressure state. At this time, the fourth gas path (7) is in a positive pressure state, the fifth gas path (8) is in a negative pressure state, the sixth gas path (9) is in a positive pressure state, and the seventh gas path (10) is in a negative pressure state. The fluid from step k1 begins to flow from the reaction outflow control channel (212) into the detection control channel (213). k3) Switch the fifth gas path (8) from negative pressure to positive pressure. At this time, the fourth gas path (7) is in positive pressure, the fifth gas path (8) is in positive pressure, the sixth gas path (9) is in positive pressure, and the seventh gas path (10) is in negative pressure. The fluid in step k2 flows from the reaction outflow control channel (212) through the detection control channel (213) into the waste liquid chamber (23).

16. A detection method, characterized in that, The application of the fluid control method as described in claim 15, wherein the reaction chamber (211) of the microfluidic device contains magnetic beads, and the microfluidic device further includes a magnetic adsorption module corresponding to the reaction chamber (211), wherein the magnetic adsorption module can turn the magnetic field on or off to achieve magnetic beads adsorbed on the bottom wall of the reaction chamber (211) or suspended in the fluid of the reaction chamber (211), wherein the surface of the magnetic beads has an anti-reflective surface, and includes the following steps: S1 Sample filling: The microfluidic device is initialized as described in step a, wherein the first reservoir (241) contains the protein to be tested and the enzyme-labeled secondary antibody solution, the second reservoir (242) contains the colorimetric solution, and the third reservoir (243) contains the stop solution. Formation of S2 magnetic bead-antibody-test protein conjugate: The pressure control module controls the first sample dispensing gas path (33) to be in a negative pressure state, and then performs the reaction chamber sample dispensing operation in step b. The test protein and enzyme-labeled secondary antibody solution in step S1 flow from the first storage chamber (241) into the reaction chamber (211) and are incubated with the magnetic beads in the reaction chamber (211) to obtain the magnetic bead-antibody-test protein conjugate mixture; S3 Magnetic Separation Waste Discharge: The magnetic adsorption module is turned on, and the magnetic bead-antibody-test protein conjugate mixture obtained in step S2 is magnetically separated into magnetic bead-antibody-test protein conjugate adsorbed onto the bottom wall of the reaction chamber (211) and the first waste liquid; the reaction chamber waste liquid discharge operation in step c is performed, and the first waste liquid flows from the reaction chamber (211) into the waste liquid chamber (23); S4 Color Development: The pressure control module controls the second sample dispensing gas path (34) to be in a negative pressure state, and then performs the reaction chamber sample dispensing operation of operation b. The color developing solution flows into the reaction chamber (211) from the second storage chamber (242). After the color developing solution has completely flowed into the reaction chamber (211), the second sample dispensing gas path (34) is controlled to be in a positive pressure state, and the magnetic adsorption module is turned off. The color developing solution reacts with the magnetic bead-antibody-test protein conjugate obtained in step S3 in the reaction chamber (211) to obtain the color developing product. S5 Termination: The pressure control module controls the third sample addition gas path (35) to be in a negative pressure state, and then performs the reaction chamber sample addition operation of operation b. The termination liquid flows into the reaction chamber (211) from the third storage chamber (243). After the termination liquid has completely flowed into the reaction chamber (211), the third sample addition gas path (35) is controlled to be in a positive pressure state. The termination liquid reacts with the colorimetric product obtained in step S4 in the reaction chamber (211) to obtain the product to be detected. S6 Detection and Analysis: Perform the operation of flowing into the detection chamber in step d. The product to be detected obtained in step S5 flows into the detection chamber (22) from the reaction chamber (211). The microfluidic device also includes a detection module for detecting the optical signal in the detection chamber. Open the detection module to detect the product to be detected in the detection chamber (22) to obtain the absorbance value. Then, take the average of the absorbance values ​​obtained from each detection channel (21) to obtain the average absorbance value, and convert it into a protein concentration value by the standard curve method.

17. The detection method according to claim 16, characterized in that, The fourth liquid storage chamber (244) stores washing liquid, and the following steps are included after step S6: S7 Washing: The pressure control module controls the fourth sample addition gas path (36) to be in a negative pressure state, and then performs the reaction chamber sample addition operation of operation b. The washing liquid flows into the reaction chamber (211) from the fourth storage chamber (244). After the stop liquid has completely flowed into the reaction chamber (211), the fourth sample addition gas path (36) is controlled to be in a positive pressure state, and the flow into the detection chamber operation of step d is performed. The washing liquid flows into the detection chamber (22) from the reaction chamber (211), and the waste liquid discharge operation of the detection chamber in step f is performed. The mixed waste liquid of the product to be detected and the washing liquid in step S6 flows into the waste liquid chamber (23) from the detection chamber (22).

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