Centrifugal microfluidic immunoassay chip

Abstract

This invention discloses a centrifugal microfluidic immunoassay chip, comprising a spiral channel. A blood chromatography chamber and a common reagent chamber are located on the side of the spiral channel near the center, while a calibration unit and several magnetic purification units for detecting different indicators are located on the side away from the center. The blood chromatography chamber is connected to the spiral channel via a paraffin valve, and the common reagent chamber is connected to the spiral channel via a paraffin valve. The end of the spiral channel is connected to a waste liquid tank. The spiral channel is connected to the calibration unit via a metering chamber, and to the magnetic purification unit via a metering chamber. This invention enables end-to-end detection, integrating the entire process of blood centrifugation to separate plasma, reagent addition and cleaning for immunoassay, and result detection on the chip. This reduces the number of detection steps, eliminating the need for sample pretreatment and post-processing of blood. The patented detection process is simple to operate, requires fewer devices, and reduces detection costs and the professional requirements for testing personnel.

Description

Technical Field

[0001] This invention pertains to detection chips, specifically a centrifugal microfluidic immunoassay chip. Background Technology

[0002] Microfluidics is a technology that uses microchannels to control and process extremely small amounts of fluid, possessing significant scientific and commercial potential. Microfluidic devices are small, low-power, and highly portable, enabling biological or chemical experiments that typically require large laboratory instruments to be performed on a chip of just a few square centimeters. Through the realization of the "Lab on a Chip" (LOC) or "Micro Total Analysis System (TAS)" concept, microfluidic technology has shown tremendous application prospects in molecular biology, analytical chemistry, and point-of-care medical diagnostics. Among these, microfluidic chips employing immunoassay techniques offer advantages such as high sensitivity and a wide calibration range.

[0003] The main techniques for immunodiagnostics include enzyme-linked immunosorbent assay (ELISA), fluorescence immunoassay, and chemiluminescence immunoassay, which have advantages such as high sensitivity and specificity. However, traditional immunoassay equipment consumes a large amount of reagents / samples, has a long testing time, and is costly, which limits the application of such products in scenarios such as entry and exit points, airports, train stations, and primary healthcare units.

[0004] In clinical diagnosis, many indicators require immunoassay for detection, and confirming a diagnosis often necessitates the simultaneous use of multiple indicators. Current standard practice involves multiple blood draws for multi-indicator testing. However, this method can lead to extreme cases of simultaneously elevated or low values ​​for certain indicators requiring combined analysis, potentially causing diagnostic errors. Furthermore, the need for multiple blood draws and lengthy waiting times for results is inconvenient for patients. Therefore, there is an urgent need to develop a testing device that can simultaneously detect multiple indicators using a small amount of blood. Summary of the Invention

[0005] Purpose of the invention: In order to overcome the shortcomings of the existing technology, the purpose of this invention is to provide a centrifugal microfluidic immunoassay chip that uses a small amount of blood, performs multi-indicator joint detection throughout the entire process, and has high accuracy.

[0006] Technical Solution: The centrifugal microfluidic immunoassay chip of the present invention includes a spiral channel. A blood chromatography chamber and a common reagent chamber are arranged on the side of the spiral channel near the center, and several magnetic purification units for detecting different indicators are arranged on the side away from the center. The blood chromatography chamber is connected to the spiral channel through a paraffin valve one, and the common reagent chamber is connected to the spiral channel through a paraffin valve two. The end of the spiral channel is connected to a waste liquid pool. The spiral channel is connected to the magnetic purification units through a metering chamber two.

[0007] Furthermore, the magnetic purification unit includes a magnetic purification reagent chamber, a paraffin valve three, a magnetic purification mixing chamber, a paraffin valve four, a primary magnetic purification capillary valve, a secondary magnetic purification capillary valve, a magnetic purification detection chamber, and a magnetic purification waste liquid chamber. The magnetic purification reagent chamber is connected to the magnetic purification mixing chamber through paraffin valve three, the metering chamber two is connected to the magnetic purification mixing chamber through the primary magnetic purification capillary valve, the magnetic purification mixing chamber is connected to the magnetic purification detection chamber through paraffin valve four, and is connected to the magnetic purification waste liquid chamber through the secondary magnetic purification capillary valve.

[0008] Furthermore, the magnetic purification reagent chamber is located near the center of the magnetic purification mixing chamber. Magnetic beads capable of switching between magnetic and non-magnetic states are placed inside the magnetic purification mixing chamber, and a magnet capable of switching between magnetic and non-magnetic states is placed at the bottom of the chamber. The magnetic beads are coated with antibodies / antigens. When the magnet is in a magnetic state, the magnetic beads are adsorbed and fixed at the bottom of the magnetic purification mixing chamber; in a non-magnetic state, the magnetic beads are suspended in the liquid within the chamber. The magnet assists in detection.

[0009] Furthermore, the difference in rotational speed between the primary magnetic purification capillary valve and the secondary magnetic purification capillary valve exceeds the threshold by more than 50 rpm. When the primary magnetic purification capillary valve is open, the secondary magnetic purification capillary valve remains closed, thereby achieving mixing of the liquid in the magnetic purification mixing chamber and realizing the order of liquid entry and exit in the magnetic purification mixing chamber.

[0010] To address the issue of consistently high sample parameters, a calibration unit is installed on the side of the spiral channel furthest from the center. The spiral channel is connected to the calibration unit via metering chamber one, which is located between the common reagent chamber and the blood chromatography chamber on the spiral channel. Since the sample used in the calibration unit is located in the standard chamber, blood is not applicable, but the common reagent is required.

[0011] Furthermore, the calibration unit includes a calibration reagent chamber, paraffin valve five, a calibration mixing chamber, paraffin valve six, a primary calibration capillary valve, a secondary calibration capillary valve, a calibration detection chamber, a calibration waste liquid chamber, a standard sample chamber, and paraffin valve seven. Metering chamber one is connected to the calibration mixing chamber through the primary calibration capillary valve; the calibration mixing chamber is connected to the calibration reagent chamber through paraffin valve five, to the standard sample chamber through paraffin valve six, to the calibration detection chamber through paraffin valve seven, and to the calibration waste liquid chamber through the secondary calibration capillary valve.

[0012] Furthermore, both the calibration reagent chamber and the standard chamber are located near the center of the calibration mixing chamber. A magnet capable of switching between magnetic and non-magnetic states is installed inside the calibration mixing chamber, and another magnet capable of switching between magnetic and non-magnetic states is located at the bottom of the calibration mixing chamber. Magnetic beads are coated with antibodies / antigens. When the magnet is in a magnetic state, the magnetic beads can be adsorbed and fixed to the bottom of the calibration mixing chamber; in a non-magnetic state, the magnetic beads are suspended in the liquid within the calibration mixing chamber.

[0013] The difference in rotational speed between the primary and secondary calibration capillary valves in the magnetic purification mixing chamber exceeds the threshold by more than 50 rpm. When the primary calibration capillary valve is open, the secondary calibration capillary valve remains closed, thereby achieving mixing of the liquid in the calibration mixing chamber and realizing the order of liquid entry and exit in the calibration mixing chamber.

[0014] Furthermore, all capillary valves and paraffin valves can be located on the back of the chip, and the paraffin valves are all controllable. Multiple common reagent chambers can be set up in proportion to the volume and sample used in each detection, and arranged along the inner side of the spiral channel.

[0015] Working Principle: Whole blood centrifugation is performed within the blood chromatography chamber. This chamber is located proximal to the chip and is equipped with a paraffin valve to prevent blood from flowing out during high-speed rotation. The paraffin valve is designed to remain closed during high-speed rotation and open during low-speed rotation. The valve opening is located anywhere on the side of the blood chromatography chamber and is directly connected to the spiral channel. After prolonged high-speed centrifugation, the whole blood will separate into two layers within the chamber: the proximal end contains plasma, and the distal end contains blood cell sediment. Reducing the rotation speed, under centrifugal force, opens the paraffin valve, allowing only the proximal plasma to pass through and enter the spiral channel.

[0016] By designing the reagent chamber, blood chromatography chamber, spiral channel, metering chamber, capillary valves, and waste liquid pool, the centrifugal force applied to the device is adjusted to allow plasma extracted from the common reagent or blood chromatography chamber to enter the spiral channel. The flow state of the liquid sample on the microfluidic chip is controlled by adjusting the rotation speed of the platform, ensuring that the sample enters and fills the metering chamber under centrifugal force, while excess liquid flows into the waste liquid pool. Then, by controlling the capillary valves (primary calibration capillary valve, secondary calibration capillary valve, primary magnetic purification capillary valve, and secondary magnetic purification capillary valve), the liquid in the metering chamber is released into the mixing chamber of the magnetic purification unit or calibration unit. The dimensions of metering chamber one and metering chamber two are designed to quantitatively control the volume entering the calibration mixing chamber and the magnetic purification mixing chamber. In this way, plasma generated from the centrifugation of the same blood sample can be included in subsequent testing processes, or liquids used in subsequent testing processes can be included in the process, achieving integrated multi-index detection within a single chip. The structure is simple and the operation is convenient.

[0017] The paraffin valve in its closed state can withstand rotational speeds up to 6000 rpm / s. The open paraffin valve has a low opening threshold, while the secondary capillary valve has a relatively high opening threshold, higher than the threshold of the open paraffin valve but lower than the threshold of the closed paraffin valve. Therefore, at low rotational speeds, with both valves closed, the liquid in the mixing chamber cannot flow out and can mix and react within the chamber. When the threshold of the secondary capillary valve is exceeded, the liquid flows to the waste chamber. When the paraffin valve is opened, the liquid flows into the detection chamber at low rotational speeds.

[0018] Open paraffin valve one and paraffin valve two to allow the public reagent or plasma or other samples obtained after blood chromatography to enter the spiral channel. Adjust the rotation speed so that the liquid enters and fills the metering chamber under centrifugal force, and excess liquid flows into the waste liquid pool. By controlling the primary magnetic purification capillary valve, the liquid in the metering chamber is released into the mixing chamber of the magnetic purification unit or calibration unit.

[0019] Instructions for using the magnetic purification mixing chamber:

[0020] a. Whole blood is injected into the blood chromatography chamber;

[0021] b. Rotate clockwise at 2000-5000 rpm for 80-600 seconds. The whole blood will separate into layers after centrifugation.

[0022] c. Open paraffin valve one, reduce the speed to 500 rpm, the plasma at the proximal end enters the spiral channel and fills all metering chambers two, and the excess plasma enters the waste liquid pool.

[0023] d. Increase the rotation speed to 600~700 rpm, the first-stage magnetic purification capillary valve opens, and the plasma in metering chamber two enters the magnetic purification mixing chamber;

[0024] e. Reduce the rotation speed to 200 rpm, and rotate clockwise and counterclockwise for 30 seconds to fully mix the blood plasma and magnetic beads;

[0025] f. Switch the magnet to magnetic state and attach the magnetic beads to the chip substrate. Rotate clockwise at 900-1000 rpm to open the secondary magnetic purification capillary valve. Unadsorbed magnetic beads and excess plasma enter the magnetic purification waste liquid chamber.

[0026] g. Open the paraffin valve two of the common reagent chamber to allow the eluent to enter the spiral channel and fill all metering chambers two. After opening the primary magnetic purification capillary valve, the eluent enters the magnetic purification mixing chamber. Switch the magnet to the non-magnetic state, and the magnetic beads suspend in the magnetic purification mixing chamber. After thoroughly cleaning by switching between clockwise and counterclockwise rotation, open the secondary magnetic purification capillary valve to allow excess eluent to enter the magnetic purification waste liquid chamber.

[0027] h. Then, open the paraffin valve three of the magnetic purification reagent chamber to allow the analyte to fully bind with the antibody / antigen used for optical detection, forming a sandwich structure.

[0028] i. After multiple cleanings and the addition of reagents, the sandwich structure is placed in a suitable substrate;

[0029] j. Open the paraffin valve four of the magnetic purification mixing chamber and rotate it clockwise at a speed of 500 rpm. The sandwich structure with the analyte and its environmental substrate enter the magnetic purification detection chamber together, and then optical detection is performed.

[0030] Instructions for calibrating the mixing chamber:

[0031] a. Open the paraffin valve seven to allow the standard in the standard chamber to enter the calibration mixing chamber;

[0032] b. Rotate clockwise and counterclockwise for 30 seconds, until the speed reaches 200 rpm, to fully mix the standard product and the magnetic beads;

[0033] c. Switch the magnet to the magnetic state and attach the magnetic beads to the chip substrate. Rotate clockwise at 900~1000 rpm. The secondary calibration capillary valve opens, and the unattached magnetic beads and excess standard enter the calibration waste liquid chamber.

[0034] d. Open the paraffin valve two of the common reagent chamber to allow the eluent to enter the spiral channel and fill the metering chamber one. After the primary calibration capillary valve is opened, the eluent enters the calibration mixing chamber. Switch the magnet to the non-magnetic state, and the magnetic beads suspend in the calibration mixing chamber. After thoroughly cleaning by switching between clockwise and counterclockwise rotation, open the secondary calibration capillary valve to allow excess eluent to enter the calibration waste liquid chamber.

[0035] e. Next, open the paraffin valve five of the calibration reagent chamber to allow the analyte to fully bind with the antibody / antigen used for optical detection, forming a sandwich structure.

[0036] f. After multiple washes and the addition of reagents, the sandwich structure is placed in a suitable substrate.

[0037] g. Open the paraffin valve seven of the calibration mixing chamber and rotate it clockwise at 500 rpm. The sandwich structure with the test sample and its environmental substrate enter the calibration detection chamber together, and then optical detection is performed.

[0038] Beneficial effects: Compared with the prior art, the present invention has the following significant features:

[0039] 1. It can realize the whole process of detection. The chip integrates the whole process of blood centrifugation to separate plasma, reagent addition and cleaning for immunoassay, and result detection. It helps to reduce the number of detection steps and eliminates the need for blood sample pretreatment and result post-processing. The patented detection process is simple, easy to operate, and requires less equipment, which reduces detection costs and the professional requirements for detection personnel.

[0040] 2. It can simultaneously detect multiple indicators. The patent divides the microfluidic chip into multiple units, each of which can detect different indicators. Blood is distributed through a spiral channel and a metering chamber. All units use the same plasma sample for detection, which can improve the accuracy of joint detection of various indicators and avoid errors of a certain indicator being too high or too low.

[0041] 3. Considering the uniformly high sample indicators, a calibration unit is set up. After calibration, the detection results of the entire microfluidic chip can be determined within a certain range, which can reduce diagnostic errors. Attached Figure Description

[0042] Figure 1 This is an exploded view of the present invention;

[0043] Figure 2 This is a top view of the present invention;

[0044] Figure 3 This is a bottom view of the present invention;

[0045] Figure 4 This is a perspective view of the present invention;

[0046] Figure 5 This is a schematic diagram of the partitioning of the present invention;

[0047] Figure 6 This is a schematic diagram showing the distribution of calibration unit 4 and magnetic purification unit 5 in this invention;

[0048] Figure 7 This is a schematic diagram of the structure of the magnetic purification unit 5 of the present invention;

[0049] Figure 8 This is a schematic diagram of the structure of the magnet 11 of the present invention;

[0050] Figure 9 This is a schematic diagram of the structure of calibration unit 4 of the present invention. Detailed Implementation

[0051] For ease of description, we use the same radius line pointing from the center of the centrifugal microfluidic immunoassay chip to the edge as a reference. The position relatively closer to the center of the chip is called the proximal end, and the position relatively farther from the center is called the distal end.

[0052] like Figure 1 Centrifugal microfluidic immunoassay chips are typically attached to a base plate 12 and a cover plate 13. Ventilation holes need to be designed on the cover plate 13, and the location and number of ventilation holes can be adjusted according to the specific application experimental scenario.

[0053] like Figures 2-6The helical channel 1 of the three-index centrifugal microfluidic immunoassay chip starts from the proximal end and extends to near the edge of the chip, with a channel at the termination end connecting to the waste liquid pool 8. Six common reagent chambers 3 and one blood chromatography chamber 2 are arranged along the proximal end of the helical channel 1, while a calibration unit 4 and three magnetic purification units 5 for different index detections are arranged at the distal end. The blood chromatography chamber 2 is connected to the helical channel 1 via paraffin valve 6, and the common reagent chamber 3 is connected to the helical channel 1 via paraffin valve 7. The helical channel 1 is connected to the calibration unit 4 via metering chamber 9, and to the magnetic purification unit 5 via metering chamber 10. The volumes of metering chambers 9 and 10 are designed as needed to allow common reagents or samples to be quantitatively introduced into different magnetic purification units 5 or calibration units 4 as required. The blood chromatography chamber 2 is located at the beginning of the helical channel 1.

[0054] Whole blood centrifugation is performed in blood chromatography chamber 2. Blood chromatography chamber 2 is located proximal to the chip and is equipped with paraffin valve 6 to prevent blood from flowing out during high-speed rotation. Paraffin valve 6 is designed to remain closed during high-speed rotation and open during low-speed rotation. The valve opening of paraffin valve 6 is located approximately halfway along the proximal edge of blood chromatography chamber 2 and is directly connected to spiral channel 1. After prolonged high-speed centrifugation, the whole blood will separate into two layers within blood chromatography chamber 2: the proximal end contains plasma separated from whole blood, and the distal end contains blood cell sediment. Reducing the rotation speed, under centrifugal force, opens paraffin valve 6, allowing only the proximal plasma to be extracted and enter spiral channel 1 through paraffin valve 6.

[0055] like Figures 7-8 The magnetic purification unit 5 includes a magnetic purification reagent chamber 501, a paraffin valve 3 502, a magnetic purification mixing chamber 503, a paraffin valve 4 504, a primary magnetic purification capillary valve 505, a secondary magnetic purification capillary valve 506, a magnetic purification detection chamber, and a magnetic purification waste liquid chamber 508. The magnetic purification reagent chamber 501 is connected to the magnetic purification mixing chamber 503 via the paraffin valve 3 502. The metering chamber 2 10 is connected to the magnetic purification mixing chamber 503 via the primary magnetic purification capillary valve 505. The magnetic purification mixing chamber 503 is connected to the magnetic purification detection chamber via the paraffin valve 4 504 and to the magnetic purification waste liquid chamber 508 via the secondary magnetic purification capillary valve 506. The magnetic purification reagent chamber 501 is located near the center of the magnetic purification mixing chamber 503. Magnetic beads 11 are installed inside the magnetic purification mixing chamber 503, and a magnet capable of switching between magnetic and non-magnetic states is installed at the bottom of the magnetic purification mixing chamber 503. Magnetic beads 11 are coated with antibodies / antigens. When magnetic beads 11 are in a magnetic state, they can be adsorbed and fixed at the bottom of the magnetic purification mixing chamber 503. When not in a magnetic state, magnetic beads 11 are suspended in the liquid in the calibration mixing chamber 403. The difference in rotational speed between the primary magnetic purification capillary valve 505 and the secondary magnetic purification capillary valve 506 exceeds the threshold value by more than 50 rpm.

[0056] like Figure 9 The calibration unit 4 includes a calibration reagent chamber 401, a paraffin valve 5 402, a calibration mixing chamber 403, a paraffin valve 6 404, a primary calibration capillary valve 405, a secondary calibration capillary valve 406, a calibration detection chamber 407, a calibration waste liquid chamber 408, a standard sample chamber 409, and a paraffin valve 7. The metering chamber 1 9 is connected to the calibration mixing chamber 403 via the primary calibration capillary valve 405. The calibration mixing chamber 403 is connected to the calibration reagent chamber 401 via paraffin valve 5 402, to the standard sample chamber 409 via paraffin valve 6 404, to the calibration detection chamber 407 via paraffin valve 7, and to the calibration waste liquid chamber 408 via the secondary calibration capillary valve 406. The calibration reagent chamber 401 and the standard sample chamber 409 are both located near the center of the calibration mixing chamber 403. A magnetic bead 11 is installed inside the calibration mixing chamber 403, and a magnet capable of switching between magnetic and non-magnetic states is installed at the bottom of the calibration mixing chamber 403. The difference in the speed exceeding the threshold between the first-stage calibration capillary valve 405 and the second-stage calibration capillary valve 406 is greater than 50 rpm.

[0057] The paraffin valves (paraffin valve 1-6, paraffin valve 2-7, paraffin valve 3-502, paraffin valve 4-504, paraffin valve 5-402, paraffin valve 6-404, paraffin valve 7) in the closed state can withstand rotational speeds up to 6000 rpm / s. The paraffin valves in the open state have a low opening threshold. The secondary capillary valves (secondary calibration capillary valve 406, secondary magnetic purification capillary valve 506) also have a relatively high opening threshold, higher than the threshold of the paraffin valves in the open state but lower than the threshold of the paraffin valves in the closed state. Therefore, at low rotational speeds, the two bottom valves are closed, preventing liquid from flowing out of the mixing chamber, allowing for mixing and reaction within the chamber. When the threshold of the secondary capillary valve is exceeded, the liquid flows to the waste liquid chamber; when the paraffin valves are opened, the liquid flows into the detection chamber at low rotational speeds.

[0058] For early gastric cancer screening, the centrifugal microfluidic immunoassay chip for the above three indicators comprises one calibration unit 4 and three magnetic purification units 5. The three magnetic purification units 5 are configured to detect G-17, PGI, and PGII, which are important serum biomarkers in early gastric cancer screening; the calibration unit 4 selects G-17 as the indicator. The specific usage method includes the following steps:

[0059] a. Whole blood is injected into blood chromatography chamber 2;

[0060] b. Rotate clockwise at 2000-5000 rpm for 80-600 seconds to separate the whole blood into layers after centrifugation;

[0061] c. Open the paraffin valve 6 and reduce the speed to 500 rpm. The plasma at the proximal end enters the spiral channel 1 and fills all the metering chambers 10. Excess plasma enters the waste liquid pool 8.

[0062] d. Increase the rotation speed to 600~700 rpm, open the first-stage magnetic purification capillary valve 505, and the plasma in metering chamber 2 10 enters the magnetic purification mixing chamber 503;

[0063] e. Reduce the rotation speed to 200 rpm, and rotate clockwise and counterclockwise for 30 seconds to fully mix the blood plasma with magnet 11;

[0064] f. Switch the magnet to the magnetic state and attract the magnetic bead 11 to the base plate 12. Rotate clockwise at a speed of 900~1000 rpm to open the secondary magnetic purification capillary valve 506. The unattracted magnet 11 and excess plasma enter the magnetic purification waste liquid chamber 508.

[0065] g. Open the paraffin valve 7 of the common reagent chamber 3 to allow the eluent to enter the spiral channel 1 and fill all metering chambers 10. After opening the primary magnetic purification capillary valve 505, the eluent enters the magnetic purification mixing chamber 503. Switch the magnet to the non-magnetic state, and the magnetic beads 11 are suspended in the magnetic purification mixing chamber 503. After thorough cleaning, open the secondary magnetic purification capillary valve 506 to allow the excess eluent to enter the magnetic purification waste liquid chamber 508.

[0066] h. Then, open the paraffin valve 502 of the magnetic purification reagent chamber 501 to allow the analyte to fully bind with the antibody / antigen used for optical detection, forming a sandwich structure.

[0067] i. After multiple washing and reagent additions, the sandwich structure is placed in a suitable substrate for chemiluminescence.

[0068] j. Open the paraffin valve 504 of the magnetic purification mixing chamber 503 and rotate it clockwise at a speed of 500 rpm. The sandwich structure with the analyte and its environmental substrate enter the magnetic purification detection chamber together, and then optical detection is performed.

[0069] If the sample indicators are uniformly high, i.e., the data is abnormal or exceeds the normal order of magnitude, then calibration unit 4 needs to be used. The method of using calibration unit 4 includes the following steps:

[0070] a. Open the paraffin valve seven to allow the standard in the standard chamber 409 to enter the calibration mixing chamber 403;

[0071] b. Rotate clockwise and counterclockwise for 30 seconds, until the speed reaches 200 rpm, to fully mix the standard product with magnetic bead 11;

[0072] c. Switch the magnet to the magnetic state and attach the magnetic bead 11 to the chip's base plate 12. Rotate clockwise at a speed of 900~1000 rpm. The secondary calibration capillary valve 406 opens, and the unattached magnetic bead 11 and excess standard enter the calibration waste liquid chamber 408.

[0073] d. Open the paraffin valve 7 of the common reagent chamber 3 to allow the eluent to enter the spiral channel 1 and fill the metering chamber 9. After the primary calibration capillary valve 405 is opened, the eluent enters the calibration mixing chamber 403. Switch the magnet to the non-magnetic state, and the magnetic bead 11 is suspended in the calibration mixing chamber 403. After thorough cleaning, open the secondary calibration capillary valve 406 to allow the excess eluent to enter the calibration waste liquid chamber 408.

[0074] e. Then, open the paraffin valve 402 of the calibration reagent chamber 401 to allow the analyte to fully bind with the antibody / antigen used for optical detection, forming a sandwich structure.

[0075] f. After multiple washes and the addition of reagents, the sandwich structure is placed in a suitable substrate.

[0076] g. Open the paraffin valve seven of the calibration mixing chamber 403 and rotate it clockwise at a speed of 500 rpm. The sandwich structure with the test sample and its environmental substrate enter the calibration detection chamber 407 together, and then optical detection is performed.

[0077] In this embodiment, calibration unit 4 can be omitted or multiple units can be set up for comparison and calibration test data.

Claims

1. A centrifugal microfluidic immunoassay chip, characterized by: The system includes a spiral channel (1), with a blood chromatography chamber (2) and a common reagent chamber (3) located on the side of the spiral channel (1) near the center, and several magnetic purification units (5) for detecting different indicators located on the side away from the center; the blood chromatography chamber (2) is connected to the spiral channel (1) via a paraffin valve (6), and the common reagent chamber (3) is connected to the spiral channel (1) via a paraffin valve (7); the end of the spiral channel (1) is connected to a waste liquid pool (8); and the spiral channel (1) is connected to the magnetic purification unit (5) via a metering chamber (10). The magnetic purification unit (5) includes a magnetic purification reagent chamber (501), a paraffin valve three (502), a magnetic purification mixing chamber (503), a paraffin valve four (504), a primary magnetic purification capillary valve (505), a secondary magnetic purification capillary valve (506), a magnetic purification detection chamber, and a magnetic purification waste liquid chamber (508). The magnetic purification reagent chamber (501) is connected to the magnetic purification mixing chamber (503) through the paraffin valve three (502). The metering chamber two (10) is connected to the magnetic purification mixing chamber (503) through the primary magnetic purification capillary valve (505). The magnetic purification mixing chamber (503) is connected to the magnetic purification detection chamber through the paraffin valve four (504) and to the magnetic purification waste liquid chamber (508) through the secondary magnetic purification capillary valve (506). A calibration unit (4) is also provided on the side of the spiral channel (1) away from the center. The spiral channel (1) is connected to the calibration unit (4) through a metering chamber (9). The metering chamber (9) is located between the common reagent chamber (3) and the blood chromatography chamber (2) on the spiral channel (1). The calibration unit (4) includes a calibration reagent chamber (401), a paraffin valve five (402), a calibration mixing chamber (403), a paraffin valve six (404), a primary calibration capillary valve (405), a secondary calibration capillary valve (406), a calibration detection chamber (407), a calibration waste liquid chamber (408), a standard sample chamber (409), and a paraffin valve seven. The metering chamber one (9) is connected to the calibration mixing chamber (403) through the primary calibration capillary valve (404). The calibration mixing chamber (403) is connected to the calibration reagent chamber (401) through the paraffin valve five (402), to the standard sample chamber (409) through the paraffin valve six (404), to the calibration detection chamber (407) through the paraffin valve seven, and to the calibration waste liquid chamber (408) through the secondary calibration capillary valve (406).

2. The centrifugal microfluidic immunoassay chip according to claim 1, wherein: The magnetic purification reagent chamber (501) is located on the near-center side of the magnetic purification mixing chamber (503).

3. The centrifugal microfluidic immunoassay chip according to claim 1, wherein: The magnetic purification mixing chamber (503) is provided with magnetic beads (11), and the bottom of the magnetic purification mixing chamber (503) is provided with a magnet that can realize the conversion between magnetic and non-magnetic states.

4. The centrifugal microfluidic immunoassay chip according to claim 1, wherein: The difference between the rotational speeds of the primary magnetic purification capillary valve (505) and the secondary magnetic purification capillary valve (506) exceeding the threshold is greater than 50 rpm.

5. The centrifugal microfluidic immunoassay chip according to claim 1, characterized in that: The calibration reagent chamber (401) and the standard chamber (409) are both located on the near-center side of the calibration mixing chamber (403).

6. The centrifugal microfluidic immunoassay chip according to claim 1, characterized in that: The calibration mixing chamber (403) is provided with a magnetic bead (11), and the bottom of the calibration mixing chamber (403) is provided with a magnet that can realize the conversion between magnetic and non-magnetic states.

7. The centrifugal microfluidic immunoassay chip according to claim 1, characterized in that: The difference between the speed limits of the primary calibration capillary valve (405) and the secondary calibration capillary valve (406) exceeds 50 rpm.

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