A microfluidic analysis chip and a sample detection method

Abstract

The present invention provides a microfluidic analysis chip and a sample detection method. The microfluidic analysis chip includes: a chip body, in which at least two microfluidic channels are arranged at the top and bottom; a blood cell detection component, including: a first micropore; an impedance signal electrode pair; a signal acquisition start electrode pair; and a signal acquisition end electrode pair. The present invention adds a sample to be tested into a microfluidic channel, and uses the impedance signal electrode pair, the signal acquisition start electrode pair, and the signal acquisition end electrode pair to collect the impedance pulse signal of the sample to be tested within a certain period of time, and detects the blood cells in the sample to be tested according to the collected impedance pulse signal. The microfluidic analysis chip is small in size, requires less samples for detection, and does not require a separate waste liquid tank. The microfluidic analysis chip of the present invention realizes one-time sample addition and obtains the detection results of blood cells and proteins at the same time, which greatly simplifies the operation steps of clinical inflammatory infection diagnosis and detection projects and reduces the sample demand.

Description

Technical Field

[0001] The present invention relates to the technical field of biological sample detection, and in particular to a microfluidic analysis chip and a sample detection method. Background Art

[0002] Biomarkers such as blood cells, hemoglobin (HGB), glycosylated hemoglobin (HbA1C), high-sensitivity C-reactive protein (hs-CRP), C-reactive protein (CRP), serum amyloid A (SAA), procalcitonin (PCT) have become routine detection items for the diagnosis of clinical inflammation-related diseases. For the above items, the existing detection technology not only realizes "separate sampling and separate detection" and "one sampling and simultaneous detection of two items", but also develops in the direction of microfluidics.

[0003] On the one hand, the existing technology has facilitated the simplification of the operation of clinical inflammatory infection diagnostic biomarker detection, but there are still problems to be solved. For example, for blood cell detection, the current detection equipment is large in size, occupies a large space, and requires a separate waste liquid tank. The waste liquid treatment is time-consuming and labor-intensive, and the reagents are multiple, which easily leads to reagent expiration and waste. At the same time, blood cell detection and inflammatory infection biomarker detection belong to two detection principles, and the required sample forms are different. For example, blood cell detection requires anticoagulated whole blood; inflammatory infection biomarker detection requires plasma or serum after removing blood cells. In actual detection, blood cells are often removed by centrifugation or filtration. In order to achieve one-time sample addition and complete the above two types of detection, the existing technology needs to integrate a blood separation device on the device. The addition of the blood separation device not only increases the complexity of the equipment and increases the failure rate of the equipment, but also increases the sample consumption as the number of detection items increases. On the other hand, the miniaturization / micro-ization in the existing technology is mainly focused on the use of a single methodology. One device can only provide simultaneous detection of different items of the same methodology, resulting in the need to configure different devices for the detection of blood cells and proteins.

[0004] Based on the current defects of blood cell testing and the inability to simultaneously detect blood cell testing and inflammatory infection biomarkers, it is necessary to make improvements. Summary of the invention

[0005] In view of this, the present invention proposes a microfluidic analysis chip and a sample detection method to solve or at least partially solve the technical problems existing in the prior art.

[0006] In a first aspect, the present invention provides a microfluidic analysis chip, comprising:

[0007] A chip body, wherein at least two microfluidic channels are disposed in the upper and lower parts thereof, and a flow channel inlet is also disposed on the chip body, and the flow channel inlet is connected to one of the microfluidic channels;

[0008] Blood cell testing kit, including:

[0009] A first micropore, which is used to connect the upper and lower microfluidic channels;

[0010] An electrical impedance signal electrode pair, which is located on both sides of the first micropore and respectively on two microchannels, and the electrical impedance signal electrode pair is used to output a blood cell electrical impedance pulse signal;

[0011] A signal acquisition start electrode pair is located in the microfluidic channel and downstream of the electrical impedance signal electrode pair, and the signal acquisition start electrode pair is used to determine the start time of blood cell electrical impedance pulse signal acquisition;

[0012] The signal acquisition end electrode pair is located in the microfluidic channel and downstream of the signal acquisition start electrode pair. The signal acquisition end electrode pair is used to determine the end time of blood cell electrical impedance pulse signal acquisition.

[0013] Preferably, the microfluidic analysis chip further comprises a protein detection component, wherein the protein detection component comprises a detection test strip located in the microfluidic channel, and the detection test strip comprises at least:

[0014] A conjugate pad coated with an antibody to the protein to be detected, the antibody having a labeled probe;

[0015] The reaction pad is located downstream of the binding pad. The reaction pad is provided with a detection line and a quality control line. The detection line is coated with antibodies of different epitopes of the protein to be detected, and the quality control line is coated with polyclonal antibodies.

[0016] Preferably, the microfluidic analysis chip has two microfluidic channels arranged in the chip body, the test strip is located in the upper microfluidic channel, the area in the upper microfluidic channel between the first micropore and the test strip is not connected, a connecting component is provided in the chip body, the connecting component includes a liquid flow signal detection electrode and a second micropore array, the second micropore array is arranged between the two microfluidic channels and is used to connect the two microfluidic channels, the lower end surface of the binding pad is fitted with the second micropore array, the liquid flow signal detection electrode is located in the lower microfluidic channel and close to the second micropore array, and the liquid flow signal detection electrode is used to detect the flow of the sample to be tested into the test strip.

[0017] Preferably, in the microfluidic analysis chip, the liquid flow signal detection electrode is in a ring shape, and the liquid flow signal detection electrode is arranged on the periphery of the second micropore array.

[0018] Preferably, in the microfluidic analysis chip, the liquid flow signal detection electrode is in the shape of a long strip and is located on one side of the second micropore array and close to the blood cell detection component.

[0019] Preferably, in the microfluidic analysis chip, the test strip further comprises a sample pad, the sample pad, the binding pad and the reaction pad are overlapped with each other in sequence, and the end surface of the sample pad is attached to the second microwell array.

[0020] Preferably, in the microfluidic analysis chip, the test strip further comprises a water absorbent pad, the water absorbent pad is located on a side of the reaction pad away from the binding pad, the water absorbent pad and the reaction pad overlap each other, the microfluidic channel where the test strip is located is provided with a flow channel outlet, and the water absorbent pad is close to the flow channel outlet.

[0021] Preferably, in the microfluidic analysis chip, an observation window is provided on the chip body corresponding to the reaction pad.

[0022] Preferably, in the microfluidic analysis chip, the protein to be tested includes at least one of CRP, SAA, PCT, HGB, hs-CRP, and HbA1c; and the labeled probe includes one of colloidal gold, fluorescent microspheres, quantum dots, organic nanoparticles, nanomagnetic materials, and lanthanide elements.

[0023] In a second aspect, the present invention further provides a sample detection method, which uses the microfluidic analysis chip for detection, and the detection method comprises:

[0024] The sample to be tested enters the microfluidic channel through the flow channel inlet;

[0025] When the sample to be tested flows to the signal collection start electrode pair, the external system starts to collect the impedance pulse signal of the blood cells through the impedance signal electrode pair, and when the sample to be tested flows to the signal collection end electrode pair, the external system stops collecting the impedance pulse signal, and the external system analyzes the sample to be tested according to the collected impedance pulse signal;

[0026] When the microfluidic analysis chip includes a protein detection component, the detection method further includes:

[0027] When the sample to be tested flows into the detection line and the quality control line, the sample to be tested reacts, and the external system analyzes the sample to be tested according to the signals generated by the detection line and the quality control line.

[0028] The microfluidic analysis chip and sample detection method of the present invention have the following beneficial effects compared with the prior art:

[0029] (1) The microfluidic analysis chip of the present invention collects the impedance pulse signal of the sample to be tested within a certain period of time by adding the sample to be tested into the microfluidic channel, using the impedance signal electrode pair, the signal collection start electrode, and the signal collection end electrode pair, and detects the blood cells in the sample to be tested according to the collected impedance pulse signal. The microfluidic analysis chip is small in size, requires less sample for detection, and does not require a separate waste liquid tank;

[0030] (2) The microfluidic analysis chip of the present invention also includes a test strip, which includes a binding pad and a reaction pad. The reaction pad is provided with a detection line and a quality control line. Protein detection is performed on the test sample according to the reaction products of the detection line and the quality control line. Through the microfluidic analysis chip, one-time sample addition is achieved, and the detection results of blood cells and proteins (such as six proteins CRP, SAA, PCT, HGB, hs-CRP, HbA1c) are obtained at the same time, which greatly simplifies the operation steps of clinical inflammatory infection diagnosis and detection projects and reduces the sample demand; at the same time, the microfluidic analysis chip of the present application realizes the combination of protein analysis detection technology and electrical impedance technology, which improves the convenience of operation. For medical staff, there is no need to prepare different types of samples (such as anticoagulant blood, procoagulant blood) or to equip multiple equipment to achieve the detection purpose, thereby reducing the equipment purchase cost. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the prior art descriptions. Obviously, the drawings described below are only some embodiments of the present invention, and for ordinary technicians in this field, other drawings can be obtained based on these drawings without creative work.

[0032] Figure 1 This is a schematic diagram of the structure of a microfluidic analysis chip in one embodiment of the present invention;

[0033] Figure 2 This is a schematic diagram of the structure of a microfluidic analysis chip in one embodiment of the present invention;

[0034] Figure 3 This is a schematic diagram of the structure of a blood cell detection component in one embodiment of the present invention;

[0035] Figure 4 This is a schematic diagram of the structure of a test strip in one embodiment of the present invention;

[0036] Figure 5 A schematic cross-sectional view of a microfluidic analysis chip having two microfluidic channels in one embodiment of the present invention;

[0037] Figure 6A schematic cross-sectional view of a microfluidic analysis chip having three microfluidic channels in one embodiment of the present invention;

[0038] Figure 7 This is a schematic diagram of the structure of a connecting component in one embodiment of the present invention;

[0039] Figure 8 This is a schematic diagram of the structure of a connecting component in one embodiment of the present invention;

[0040] Fig. 9 A schematic cross-sectional view of a microfluidic analysis chip having four microfluidic channels in one embodiment of the present invention;

[0041] Fig.10 This is a schematic diagram of the structure of a microfluidic analysis chip in one embodiment of the present invention;

[0042] Fig.11 A working timing diagram of a microfluidic analysis chip in one embodiment of the present invention when performing detection;

[0043] Fig.12 A timing diagram of the operation of the microfluidic analysis chip during detection in one embodiment of the present invention. DETAILED DESCRIPTION

[0044] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0045] The present application embodiment provides a microfluidic analysis chip, such as Figures 1 to 3 As shown, including:

[0046] The chip body 1 has at least two microfluidic channels 2 disposed therein, and a channel inlet 11 is also disposed on the chip body 1, and the channel inlet 11 is connected to one of the microfluidic channels 2;

[0047] Blood cell detection component 3, comprising:

[0048] A first micropore 31, which is used to connect the upper and lower microfluidic channels 2;

[0049] The electrical impedance signal electrode pair 32 is located at both sides of the first microhole 31 and respectively located on the two microchannels 2. The electrical impedance signal electrode pair 32 is used to output a blood cell electrical impedance pulse signal;

[0050] The signal acquisition start electrode pair 33 is located in the microfluidic channel 2 and downstream of the electrical impedance signal electrode pair 32. The signal acquisition start electrode pair 33 is used to determine the start time of blood cell electrical impedance pulse signal acquisition;

[0051] The signal acquisition end electrode pair 34 is located in the microfluidic channel 2 and downstream of the signal acquisition start electrode pair 33. The signal acquisition end electrode pair 34 is used to determine the end time of blood cell electrical impedance pulse signal acquisition.

[0052] It should be noted that the microfluidic analysis chip in the embodiment of the present application can work in both a positive pressure environment and a negative pressure environment; at least two microfluidic channels 2 are arranged from top to bottom in the chip body 1. Specifically, the number of microfluidic channels 2 can be determined according to actual usage, such as 2, 3, 4, 5, etc.; since the multiple microfluidic channels 2 are interconnected, the multiple microfluidic channels 2 essentially constitute a complete microfluidic channel. For example, two microfluidic channels 2 can be arranged in the chip body 1. In order to facilitate preparation and processing, the chip body 1 can be set as a three-layer structure, and the microfluidic channels 2 are respectively arranged on the upper and lower surfaces of the middle layer, and the upper and lower layers are respectively attached to the middle layer. The channel inlet 11 is opened in the upper layer, and the chip body 1 can be formed; refer to Figures 1-2 As shown, Figures 1-2 The microfluidic channels 2 arranged on the upper and lower surfaces of the middle layer in different schemes, the flow direction of the sample to be tested (the direction of the arrow in the figure) and the setting position of the blood cell detection component 3 ( Figure 1 The test strip is located in the microfluidic channel where the upper surface is located. Figure 2 The test strip is located in the microfluidic channel where the lower surface is located); a flow channel inlet 11 is opened on the chip body 1, and the sample to be tested can be added to the microfluidic channel 2 through the flow channel inlet 11. At the same time, the flow channel inlet 11 provides an interface for an external power source (when working in a positive pressure environment); a blood cell detection component 3 is set in the microfluidic channel 2 to detect blood cells in the sample. For details, refer to Figure 3 As shown, the blood cell detection component 3 includes a first micropore 31, an impedance signal electrode pair 32, a signal acquisition start electrode pair 33, and a signal acquisition end electrode pair 34, wherein the first micropore 31 is used to connect the upper and lower microfluidic channels 2. Specifically, the aperture of the first micropore is 30-60 μm, and the impedance signal electrode pair 32 is located on both sides of the first micropore 31 and is respectively located on the upper and lower microfluidic channels 2, that is, one of the impedance signal electrode pairs 32 is located on the upper microfluidic channel 2, and the other impedance signal electrode pair 32 is located on the lower microfluidic channel 2; the signal acquisition start electrode pair 33 is located downstream of the impedance signal electrode pair 32, and the signal acquisition end electrode pair 34 is located downstream of the signal acquisition start electrode pair 33. It should be pointed out that the downstream in the present application is the flow direction of the sample to be tested in the microfluidic channel ( Figures 1-2The upstream and downstream directions are described by using the arrow direction in the middle. When the sample to be tested flows into the first micropore 31 through the microfluidic channel 2 above, when the blood cells in the sample to be tested pass through the first micropore 31, the blood cells replace the same volume of liquid in the first micropore 31, and due to the volume replacement effect and the difference in conductivity between the blood cells and the surrounding wrapping medium, an impedance pulse signal is generated, and the impedance signal electrode pair 32 can output the generated impedance pulse signal; the signal acquisition start electrode pair 33 determines the start time of the blood cell impedance signal acquisition; the signal acquisition end electrode pair 34 determines the end time of the blood cell impedance signal acquisition; the signal acquisition start electrode pair 33 and the signal acquisition end electrode pair 34 are combined to determine the volume of the sample solution to be tested whose impedance pulse signal is collected. After the sample to be tested flows through the impedance signal electrode pair 32, the signal acquisition start electrode pair 33, and the signal acquisition end electrode pair 34, blood cell detection is performed. In practice, an external system is also included, and the external system is specifically a host computer. The impedance signal electrode pair 32, the signal acquisition start electrode pair 33, and the signal acquisition end electrode pair 34 are all electrically connected to the external system. The external system collects the impedance pulse signal from the impedance signal electrode pair 32 from the start time to the end time. The external system can be used to classify and count the red blood cell / platelet combination and a type of blood cell in the white blood cell according to the blood sample processing method to be tested, so as to detect the concentration parameters of the corresponding blood cells / blood cell subpopulations. The microfluidic analysis chip of the present application, by adding the sample to be tested into the microfluidic channel, uses the impedance signal electrode pair, the signal acquisition start electrode, and the signal acquisition end electrode pair to collect the impedance pulse signal of the sample to be tested within a certain period of time, and detects the blood cells in the sample to be tested according to the collected impedance pulse signal. The microfluidic analysis chip is small in size, requires less samples for detection, and does not require a separate waste liquid tank.

[0053] In some embodiments, Figure 4 As shown, it also includes a protein detection component, which includes a detection test strip 4 located in the microfluidic channel 2, and the detection test strip 4 at least includes:

[0054] A conjugate pad 41, which is coated with an antibody to be detected protein, and the antibody is provided with a labeled probe;

[0055] The reaction pad 42 is located downstream of the binding pad 41. A detection line 43 and a quality control line 44 are arranged on the reaction pad 42. The detection line 43 is coated with antibodies of different epitopes of the protein to be detected, and the quality control line 44 is coated with polyclonal antibodies.

[0056] In the above embodiment, the test strip 4 is located in the microfluidic channel 2 and downstream of the blood cell detection component 3. Specifically, two microfluidic channels 2 are used for illustration. The two microfluidic channels 2 flow through the first micropore 31, and the test strip 4 is located in the lower microfluidic channel 2. The channel inlet 11 is connected to the upper microfluidic channel 2. The sample to be tested enters the upper microfluidic channel 2 through the channel inlet 11 and then passes through the first micropore 31. At this time, the detection of blood cells in the sample is completed; then the sample continues to flow to the test strip 4 in the lower microfluidic channel 2 for protein detection. A plurality of detection lines 43 may be provided on the reaction pad 42, and the detection line 43 is located upstream of the quality control line 44. When the sample to be tested contains one or more antigens to be tested, the antigen combines with the test protein antibody of the labeled probe to form a "labeled probe-antibody-antigen" complex; when the sample to be tested flows through the reaction pad 42, the complex combined with the antigen forms a "labeled probe-antibody-antigen-antibody" complex at the position of the detection line 43, and is fixed at the position of the detection line to form a reaction signal; the "labeled probe-antibody" complex not combined with the antigen forms a "labeled probe-antibody-antibody complex" at the position of the quality control line 44, and is fixed at the position of the quality control line to form a reference signal. According to the reaction signal and the reference signal, the protein in the sample to be tested can be detected and analyzed. In the above technical scheme, one-time sampling is achieved, and the test results of blood cells and proteins (such as six proteins CRP, SAA, PCT, HGB, HbA1C, and hs-CRP) are obtained at the same time, which greatly simplifies the operation steps of clinical inflammatory infection diagnosis and detection projects and reduces the sample demand; at the same time, the microfluidic analysis chip of the present application realizes the combination of protein analysis and detection technology and electrical impedance technology, which improves the convenience of operation. For medical staff, there is no need to prepare different types of samples (such as anticoagulants and procoagulants), nor to equip multiple devices to achieve the purpose of detection, which reduces the cost of equipment purchase and can significantly reduce the integration of the external system supporting it. The liquids are encapsulated inside the chip and are disposable. The combination of the two technologies lays the foundation for the development of low failure rate and multi-parameter equipment.

[0057] In some embodiments, two microfluidic channels 2 are provided in the chip body 1, and the test strip 4 is located in the upper microfluidic channel 2. The area between the first micropore 31 and the test strip 4 in the upper microfluidic channel 2 is not connected. A connecting component 5 is provided in the chip body 1. The connecting component 5 includes a liquid flow signal detection electrode 51 and a second micropore array 52. ​​The second micropore array 52 is arranged between the two microfluidic channels 2 and is used to connect the two microfluidic channels 2. The lower end surface of the binding pad 41 is in contact with the second micropore array 52. ​​The liquid flow signal detection electrode 51 is located in the lower microfluidic channel 2 and is close to the second micropore array 52. ​​The liquid flow signal detection electrode 51 is used to detect the flow of the sample to be tested into the test strip 4.

[0058] In the above embodiment, the test strip 4 is located in the upper microfluidic channel 2. Figure 5 As shown, a partition 21 is provided in the upper microfluidic channel 2 to prevent the area between the first micropore 31 and the test paper strip 4 in the microfluidic channel 2 from being connected. The sample to be tested flows into the upper microfluidic channel 2 through the flow channel inlet 11, flows into the lower microfluidic channel 2 through the first micropore 31, and then flows into the lower end surface of the binding pad 41 of the test paper strip 4 through the second micropore array 52. ​​The diameter of the second micropore in the second micropore array 52 is 60 to 100 μm, and the protein in the sample to be tested is then detected. At the same time, a liquid flow signal detection electrode 51 is provided in the lower microfluidic channel 2 and near the second micropore array 52. ​​The sample to be tested flows through the liquid flow signal detection electrode 51 and then flows out through the second micropore array 52. ​​The liquid flow signal detection electrode 51 can detect that the sample to be tested flows into the test paper strip 4.

[0059] In some embodiments, Figure 6 As shown, three microfluidic channels 2 are arranged in the chip body 1, and the two upper microfluidic channels 2 are connected through the first micropore 31, that is, the uppermost and middle two microfluidic channels 2 are connected through the first micropore 31. Obviously, the blood cell detection can be completed by the sample to be tested through the first micropore 31; a second micropore array 52 is provided between the two microfluidic channels 2 at the bottom and the middle in the chip body 1, and the two microfluidic channels 2 at the bottom and the middle can be connected through the second micropore array 52; the test strip 4 is located at the bottom micropore The sample to be tested flows into the middle microfluidic channel and then flows into the surface of the conjugated pad 41 through the second micropore array 52, thereby detecting the protein in the sample to be tested. Meanwhile, a liquid flow signal detection electrode 51 is arranged in the middle microfluidic channel 2 and close to the second micropore array 52. ​​The sample to be tested flows through the liquid flow signal detection electrode 51 and then flows out through the second micropore array 52. ​​The liquid flow signal detection electrode 51 can detect that the sample to be tested flows into the test paper strip 4.

[0060] In some embodiments, Figure 7 As shown, the liquid flow signal detection electrode 51 is in a ring shape and is disposed at the periphery of the second micropore array 52 , that is, the second micropore array 52 is disposed in the central hole of the liquid flow signal detection electrode 51 .

[0061] In some embodiments, Figure 8 As shown, the liquid flow signal detection electrode 51 is in a strip shape, and the liquid flow signal detection electrode 51 is located in the second micropore array 52 and close to one side of the blood cell detection component 3 .

[0062] It should be pointed out that the liquid flow signal detection electrode 51 of the present application can be in any shape as long as it can realize the function of detecting the flow of the sample to be tested into the test paper strip 4. The above-mentioned circular ring shape and long strip shape are only two preferred shapes.

[0063] In some embodiments, four microfluidic channels 2 are provided in the chip body 1. Fig. 9 As shown, for the convenience of description, they are named as the first microfluidic channel, the second microfluidic channel, the third microfluidic channel, and the fourth microfluidic channel from top to bottom, wherein the first microfluidic channel and the second microfluidic channel are connected through the first micropore 31, and the second microfluidic channel and the fourth microfluidic channel are connected through the first micropore 31; and the above-mentioned second micropore array 52 is arranged between the third microfluidic channel and the fourth microfluidic channel, and the test strip 4 is located in the third microfluidic channel, and the lower end surface of the conjugate pad 41 is attached to the second micropore array 52. ​​In this solution, the sample to be tested passes through the first micropore 3 1 enters the second microfluidic channel, then passes through the first micropore 31 to enter the fourth microfluidic channel, and then passes through the second micropore array 52 to flow upward to the test paper strip 4 of the third microfluidic channel to perform protein detection on the sample to be tested; obviously, a liquid flow signal detection electrode 51 can also be set in the fourth microfluidic channel near the second micropore array 52. ​​After the sample to be tested flows through the liquid flow signal detection electrode 51, it passes through the second micropore array 52 and flows into the test paper strip 4 of the third microfluidic channel. The liquid flow signal detection electrode 51 can detect that the sample to be tested flows into the test paper strip 4.

[0064] In some embodiments, the test strip 4 further includes a sample pad 45 . The sample pad 45 , the binding pad 41 , and the reaction pad 42 are overlapped with each other in sequence. The upper end surface of the sample pad 45 is attached to the second microwell array 52 .

[0065] In the above embodiment, after the sample flows into the sample pad, it continues to flow forward to the binding pad 41 and the reaction pad 42 to react.

[0066] In some embodiments, please refer again to Figure 4 As shown, the test strip 4 also includes a water absorbent pad 46, which is located on the side of the reaction pad 42 away from the binding pad 41. The water absorbent pad 46 and the reaction pad 42 are overlapped or integrally formed with each other. The microfluidic channel 2 where the test strip 4 is located is provided with a flow channel outlet 12, and the water absorbent pad 46 is close to the flow channel outlet 12.

[0067] In the above embodiment, the test strip 4 includes a sample pad 45, a binding pad 41, a reaction pad 42 and a water-absorbing pad 46 which are overlapped with each other. The water-absorbing pad 46 is made of absorbent paper. The sample to be tested flows through the sample pad 45, the binding pad 41, the reaction pad 42 and the water-absorbing pad 46 in sequence. In practice, when the microfluidic analysis chip works in a positive pressure environment, the flow channel outlet 12 is directly connected to the atmosphere; when the microfluidic analysis chip works in a negative pressure environment, the flow channel outlet 12 is connected to an external negative pressure power source. During operation, the sample to be tested is added to the microfluidic channel 2 through the flow channel inlet 11, and under the action of the external power source or the capillary force of the microfluidic channel 2, it flows through the blood cell detection component 3 to complete the blood cell detection in the sample to be tested, and flows through the test strip 4 to complete the protein detection.

[0068] In some embodiments, Fig.10 As shown, an observation window 13 is provided on the chip body 1 corresponding to the reaction pad 42 .

[0069] In the above embodiment, Fig.10 The figure shows the microfluidic channels 2 arranged on the upper and lower surfaces in another scheme, the flow direction of the sample to be tested (the direction of the arrow in the figure), and the setting positions of the blood cell detection component 3 and the test strip 4. Specifically, the chip body 1 is a flat layered three-dimensional structure, and the chip body 1 can be made of a transparent material, such as polymethyl methacrylate. The chip body 1 made of a transparent material can realize the observation of the complex formed by the sample to be tested at the detection line 43 and the complex formed at the quality control line 44.

[0070] In some embodiments, the protein to be tested includes at least one of CRP, SAA, PCT, HGB, hs-CRP, and HbA1C; the labeled probe includes one of colloidal gold, fluorescent microspheres, quantum dots, organic nanoparticles, nanomagnetic materials, and lanthanide elements. Preferably, the labeled probe is a quantum dot.

[0071] Based on the same inventive concept, the present application also provides a sample detection method, which uses the above-mentioned microfluidic analysis chip for detection. The detection method specifically includes:

[0072] The sample to be tested enters the microfluidic channel through the flow channel inlet;

[0073] When the sample to be tested flows to the signal collection start electrode pair, the external system starts to collect the impedance pulse signal of the blood cells through the impedance signal electrode pair, and when the sample to be tested flows to the signal collection end electrode pair, the external system stops collecting the impedance pulse signal, and the external system analyzes the sample to be tested according to the collected impedance pulse signal;

[0074] When the microfluidic analysis chip includes a protein detection component, the detection method further includes:

[0075] When the sample to be tested flows into the detection line and the quality control line, the sample to be tested reacts, and the external system analyzes the sample to be tested according to the signals generated by the detection line and the quality control line.

[0076] In some embodiments, if the microfluidic analysis chip includes a liquid flow signal detection electrode and a second micropore array, the detection method includes: after the sample to be tested flows through the signal acquisition end electrode pair, it flows into the binding pad and the reaction pad through the liquid flow signal detection electrode and the second micropore array, the sample to be tested reacts, and the sample to be tested is analyzed according to the reaction products of the detection line and the quality control line.

[0077] Specifically, Fig.11 As shown, the working timing diagram of the microfluidic analysis chip of the present application when performing detection. At t0, the sample to be tested is added to the flow channel entrance, the external power source starts to work (high level), or the capillary force starts to work, or the external power source and the capillary force work together; at t1, the sample to be tested flows to the signal acquisition start electrode (low level), and the external system connected to the microfluidic analysis chip starts the acquisition of the impedance signal; at t2, the sample to be tested flows to the signal acquisition end electrode (low level), and the external system connected to the microfluidic analysis chip ends the acquisition of the impedance signal; at t3, all the samples to be tested flow through the signal acquisition end electrode, and the external power source stops working (low level). Subsequently, wait for T minutes, the sample to be tested flows all reach the absorbent pad, and the external system connected to the microfluidic analysis chip uses the observation window to detect the signals generated at the detection line and the quality control line position. It should be pointed out that in the above working sequence, since the microfluidic analysis chip does not contain a liquid flow signal detection electrode, at this time, the signal acquisition end electrode also takes into account the function of the liquid flow signal detection electrode, that is, the signal acquisition end electrode is used to detect the flow of the sample to be tested.

[0078] Another working sequence diagram of the microfluidic analysis chip of the present application when performing detection, such as Fig.12As shown. At t0, the sample to be tested is added to the flow channel inlet 1, the external power source starts to work (high level), or the capillary force starts to work, or the external power source and the capillary force work together; at t1, the sample flow to be tested reaches the signal acquisition start electrode (low level), and the external system connected to the microfluidic analysis chip starts to collect the electrical impedance signal; at t2, the sample flow to be tested reaches the signal acquisition end electrode (low level), and the external system connected to the blood cell analysis chip ends the collection of the electrical impedance signal; at t3, the sample flow to be tested reaches the liquid flow signal detection electrode, and the liquid flow signal detection electrode turns to a low level; at t4, the sample flow to be tested flows through the liquid flow signal detection electrode, the liquid flow signal detection electrode turns to a high level, and the external power source stops working (low level); then, after waiting for T minutes, the sample flow to be tested reaches the water absorption pad, and the external system connected to the microfluidic analysis chip uses the observation window to detect the signals generated at the detection line and the quality control line. After the test is completed, the external system connected to the microfluidic analysis chip calculates the concentration parameters of the blood cells / blood cell subpopulations to be tested and the protein to be tested according to the collected impedance pulse signal, the detection line, and the signal strength at the quality control line position, and outputs a test report, realizing one-time sampling and reporting the test results of biomarkers such as the blood cells to be tested and the protein to be tested required for the diagnosis of clinical inflammatory infections, greatly simplifying the clinical inflammatory infection detection operation steps.

[0079] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. A microfluidic analysis chip, characterized in that: include: A chip body, wherein at least two microfluidic channels are disposed in the upper and lower parts thereof, and a flow channel inlet is also disposed on the chip body, and the flow channel inlet is connected to one of the microfluidic channels; Blood cell testing kit, including: A first micropore, which is used to connect the upper and lower microfluidic channels; An electrical impedance signal electrode pair, which is located on both sides of the first micropore and respectively on two microchannels, and the electrical impedance signal electrode pair is used to output a blood cell electrical impedance pulse signal; A signal acquisition start electrode pair is located in the microfluidic channel and downstream of the electrical impedance signal electrode pair, and the signal acquisition start electrode pair is used to determine the start time of blood cell electrical impedance pulse signal acquisition; A signal acquisition end electrode pair is located in the microfluidic channel and downstream of the signal acquisition start electrode pair, and the signal acquisition end electrode pair is used to determine the end time of blood cell electrical impedance pulse signal acquisition; A protein detection component, the protein detection component comprising a detection test strip located in a microfluidic channel, the detection test strip comprising at least: A conjugate pad coated with an antibody to the protein to be detected, the antibody having a labeled probe; A reaction pad is located downstream of the binding pad, and a detection line and a quality control line are arranged on the reaction pad. The detection line is coated with antibodies of different epitopes of the protein to be detected, and the quality control line is coated with polyclonal antibodies; Three microfluidic channels are arranged in the chip body, the two upper microfluidic channels are connected through a first micropore, that is, the uppermost and middle two microfluidic channels are connected through the first micropore, and the sample to be tested can complete blood cell detection through the first micropore; a second micropore array is provided between the two microfluidic channels located at the bottom and the middle in the chip body, and the two microfluidic channels at the bottom and the middle can be connected through the second micropore array; the test paper strip is located in the lowermost microfluidic channel, and the upper end surface of the binding pad is fitted with the second micropore array, and the sample to be tested flows into the middle microfluidic channel and then flows into the surface of the binding pad through the second micropore array, and then the protein in the sample to be tested is detected; at the same time, a liquid flow signal detection electrode is arranged in the middle microfluidic channel and close to the second micropore array, and the sample to be tested flows through the liquid flow signal detection electrode and then flows out through the second micropore array, and the liquid flow signal detection electrode can detect that the sample to be tested flows into the test paper strip.

2. The microfluidic analysis chip according to claim 1, characterized in that: The liquid flow signal detection electrode is in a ring shape and is arranged on the periphery of the second micropore array.

3. The microfluidic analysis chip according to claim 1, characterized in that: The liquid flow signal detection electrode is in the shape of a long strip and is located at one side of the second micropore array and close to the blood cell detection component.

4. The microfluidic analysis chip according to claim 1, characterized in that: The test strip further comprises a sample pad. The sample pad, the binding pad and the reaction pad are overlapped with each other in sequence, and the upper end surface of the sample pad is attached to the second microwell array.

5. The microfluidic analysis chip according to claim 4, characterized in that: The test strip also includes a water absorbent pad, which is located on the side of the reaction pad away from the binding pad. The water absorbent pad overlaps the reaction pad. The microfluidic channel where the test strip is located is provided with a flow channel outlet, and the water absorbent pad is close to the flow channel outlet.

6. The microfluidic analysis chip according to claim 1, characterized in that: An observation window is provided on the chip body at a location corresponding to the reaction pad.

7. The microfluidic analysis chip according to claim 1, characterized in that: The protein to be tested includes at least one of CRP, SAA, PCT, HGB, hs-CRP, and HbA1c; the labeling probe includes one of colloidal gold, fluorescent microspheres, quantum dots, organic nanoparticles, nanomagnetic materials, and lanthanide elements.

8. A sample detection method, characterized in that: The microfluidic analysis chip according to any one of claims 1 to 7 is used for detection, wherein the detection method comprises: The sample to be tested enters the microfluidic channel through the flow channel inlet; When the sample to be tested flows to the signal collection start electrode pair, the external system starts to collect the impedance pulse signal of the blood cells through the impedance signal electrode pair, and when the sample to be tested flows to the signal collection end electrode pair, the external system stops collecting the impedance pulse signal, and the external system analyzes the sample to be tested according to the collected impedance pulse signal; When the microfluidic analysis chip includes a protein detection component, the detection method further includes: When the sample to be tested flows into the detection line and the quality control line, the sample to be tested reacts, and the external system analyzes the sample to be tested according to the signals generated by the detection line and the quality control line.

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