Kit for typing detection of red blood cell IgG antibody by chemiluminescence immunoassay

Abstract

The invention relates to the technical field of biological detection, in particular to a kit for typing detection of a red blood cell IgG antibody by a chemiluminescence immunoassay method. The invention relates to a kit for typing detection of red blood cell IgG antibodies by chemiluminescence immunoassay. The kit comprises a reagent R1, magnetic bead particles coated with different anti-human IgG subtype antibodies and a preserving fluid I of the magnetic bead particles, a reagent R2, an acridinium ester-labeled anti-human IgG antibody and a preservation solution II thereof; a reagent R3; a reagent R4; pre-excitation liquid and excitation liquid; wherein the concentration of the magnetic bead particles coated with different anti-human IgG subtype antibodies in the preserving fluid I is 0.3-0.4 mg / ml, and the concentration of the acridinium ester-labeled anti-human IgG antibodies in the preserving fluid II is 95-100 ng / ml. The kit can simultaneously detect four IgG subtypes on red blood cells, provides comprehensive basis for AIHA disease diagnosis, is matched with an automatic analyzer for use, reduces errors, improves the detection accuracy, and improves the detection accuracy. And the preparation process is efficient and is suitable for large-scale production.

Description

Technical Field

[0001] The present application relates to the field of biological detection technology, and in particular to a kit for detecting erythrocyte IgG antibodies by chemiluminescent immunoassay typing. Background Art

[0002] In recent years, with the in-depth study of autoimmune hemolytic anemia (AIHA), especially the analysis of IgG antibody subtypes on red blood cells, it has become the key to understanding and treating this complex disease. At present, the clinical diagnosis of AIHA mainly relies on the direct antiglobulin test (DAT), which can detect incomplete antibodies and / or complement on the surface of red blood cells, providing an important basis for the diagnosis of AIHA.

[0003] However, traditional DAT tests can often only provide qualitative results during the detection process, that is, whether there are incomplete antibodies or complement on red blood cells, but cannot distinguish the specific types of antibodies or complement, which poses a challenge to further understanding the pathogenesis of the disease and formulating targeted treatment plans.

[0004] In IgG immune hemolytic anemia, different IgG subtypes differ in their ability to activate complement and bind to the Fc receptors of phagocytes, so their effects on red blood cell destruction are also different. IgG1 and IgG3 are often closely related to the acceleration of red blood cell destruction and the severity of anemia in the acute phase of the disease, especially IgG3, which usually causes more severe red blood cell destruction.

[0005] Therefore, the development of a kit that can accurately detect IgG antibody subtypes on red blood cells is of great significance for improving the diagnostic accuracy of AIHA, assessing the degree of anemia, and predicting the treatment effect. Summary of the invention

[0006] In view of the deficiency of the direct antiglobulin test (DAT) in the prior art that it is unable to accurately detect the IgG antibody subtype on red blood cells, the present application provides a chemiluminescent immunoassay typing and detection kit for red blood cell IgG antibodies and a preparation method thereof. The kit can be used in conjunction with a fully automatic chemiluminescent immunoassay instrument to efficiently and accurately detect four subtypes of antibodies, IgG1, IgG2, IgG3 and IgG4, on red blood cells, thereby providing a more accurate basis for the diagnosis of autoimmune hemolytic anemia (AIHA), the assessment of the degree of anemia and the formulation of treatment plans.

[0007] The present application provides a chemiluminescent immunoassay method for typing and detecting red blood cell IgG antibodies using the following technical solution: A kit for typing and detecting red blood cell IgG antibodies by chemiluminescent immunoassay, the kit comprising: reagent R1, magnetic beads coated with different anti-human IgG subtype antibodies and preservation solution I; reagent R2, acridinium ester-labeled anti-human IgG antibodies and preservation solution II; reagent R3; reagent R4; pre-excitation solution and excitation solution; wherein the concentration of the magnetic beads coated with different anti-human IgG subtype antibodies in preservation solution I is 0.3-0.4 mg / ml, and the concentration of the acridinium ester-labeled anti-human IgG antibodies in preservation solution II is 95-100 ng / ml.

[0008] By adopting the above technical solution: it is possible to simultaneously detect four subtypes of antibodies, namely IgG1, IgG2, IgG3, and IgG4, providing a more comprehensive and accurate basis for the diagnosis, disease assessment, and treatment plan formulation of diseases such as autoimmune hemolytic anemia (AIHA). In addition, when used in conjunction with a fully automatic chemiluminescence immunoassay analyzer, the automation and standardization of the detection process is achieved, human operating errors are reduced, and detection efficiency and repeatability are improved.

[0009] The reasons may be as follows: the kit uses specific antibodies for different IgG subtypes coated on magnetic beads. This highly specific binding can accurately capture the target IgG antibody subtype in the sample and reduce the background noise caused by non-specific binding. Acridinium ester is used as a luminescent marker, which has the advantages of high sensitivity, good stability, and high luminescence intensity. It can produce strong chemiluminescent signals under excitation conditions, thereby improving the sensitivity of detection; The optimized magnetic bead concentration (0.3-0.4 mg / ml) ensures sufficient capture area and stability. It is neither too dense to reduce capture efficiency nor too sparse to affect detection sensitivity. The concentration of acridinium ester-labeled anti-human IgG antibody (95-100 ng / ml) was also carefully optimized to achieve the best balance between signal intensity and detection sensitivity; When used in conjunction with a fully automatic chemiluminescence immunoassay analyzer, a fully automated process from sample processing to result output is realized, and the four main subtypes of IgG (IgG1, IgG2, IgG3, and IgG4) can be detected simultaneously, which not only improves detection efficiency, but also reduces errors caused by human operation, ensuring the accuracy and repeatability of test results. This comprehensive detection capability provides richer information for clinicians, helping doctors to more accurately judge the type of disease, the severity of the disease, and the comprehensive coverage of prognosis.

[0010] Preferably, the volume ratio of reagent R1, reagent R2, reagent R3, and reagent R4 is 1:(2-2.6):(2-4):(2-4).

[0011] By adopting the above technical solution, the detection sensitivity and accuracy of the chemiluminescent immunoassay typing and detection kit for red blood cell IgG antibodies can be improved, while maintaining the stability and repeatability of the reaction system.

[0012] Preferably, the different anti-human IgG subtype antibodies include anti-human IgG1 subtype antibodies, anti-human IgG2 subtype antibodies, anti-human IgG3 subtype antibodies, and anti-human IgG4 subtype antibodies.

[0013] By adopting the above technical solution: the accuracy of the test can be improved. In autoimmune diseases such as autoimmune hemolytic anemia (AIHA), the increase of specific IgG subtype antibodies may have different clinical evaluation significance. Therefore, the detection of these subtype antibodies can improve the accuracy and specificity of the test.

[0014] Preferably, the preparation steps of the kit include: a. Preparation of reagent R1: prepare magnetic beads coated with different anti-human IgG subtype antibodies and preservation solution I respectively, then evenly select different types of magnetic beads coated with anti-human IgG subtype antibodies, mix them evenly and put them into preservation solution I, and dilute the anti-IgG subtype antibodies coated with magnetic beads with preservation solution I to 0.4 mg / ml, that is; b. Preparation of reagent R2: prepare acridinium ester-labeled anti-human IgG antibody and preservation solution II respectively, then place the acridinium ester-labeled anti-human IgG antibody into preservation solution II, and dilute the acridinium ester-labeled anti-human IgG antibody to 100 ng / ml with preservation solution II; c. Preparation of reagent R3: Weigh Tris, sodium chloride, and BSA, put them into a beaker, dissolve the solids with ultrapure water, and transfer them to a centrifuge tube to obtain; d. Preparation of reagent R4: Weigh Tris, sodium chloride, and BSA, put them into a beaker, dissolve the solids with ultrapure water, and transfer them to a centrifuge tube.

[0015] By adopting the above technical solution: accurately controlling the composition and concentration of each reagent, the quality and performance of the chemiluminescent immunoassay typing and detection kit for red blood cell IgG antibodies are ensured.

[0016] Preferably, the method for preparing magnetic beads coated with anti-human IgG subtype antibodies comprises: taking magnetic beads, removing the supernatant after vortex separation, washing, adding anti-human IgG antibodies and dispersing them in a coating buffer, mixing and incubating; After the incubation is completed, the supernatant is removed, the magnetic beads are washed, and redispersed in the preservation solution I. After blocking and incubation, the result is obtained.

[0017] By adopting the above technical solution: the method has clear steps, simple operation, and is suitable for large-scale production and application. By relying on vortex separation, washing, incubation and blocking steps, high-quality coated antibody magnetic beads can be efficiently prepared, providing a powerful tool for subsequent immunoassay and detection.

[0018] Preferably, the preparation method of the acridinium ester-labeled anti-human IgG antibody is as follows: The anti-human IgG antibody to be labeled and acridinium ester are mixed and dispersed in a labeling buffer, wrapped with tin foil, and allowed to react in a constant temperature and humidity chamber at 35-37°C for 0.5-1h to obtain an anti-human IgG antibody mixture labeled with acridinium ester; Subsequently, the obtained acridinium ester-labeled anti-human IgG antibody mixture is placed in an ultrafiltration tube for ultrafiltration to remove unreacted acridinium ester, thereby obtaining the acridinium ester-labeled anti-human IgG antibody.

[0019] By adopting the above technical solution: efficient, stable and high-purity labeled antibodies can be obtained.

[0020] Preferably, the preservation solution I is prepared by the following preparation method: Weigh 3g Tris and 8g NaCl into a beaker, and pour in 800ml ultrapure water, stir; adjust the pH of the solution in the beaker to 8.0 with 1M HCl solution; weigh 3g BSA and add it to the solution, and measure 1mL Proclin-300, add it to the solution, mix well and make up to 1000ml.

[0021] By adopting the above technical solution: the preservation solution I provides a stable and suitable storage environment for the anti-human IgG antibodies coated on the magnetic beads, which helps to maintain the activity and stability of the antibodies, reduce nonspecific binding, and reduce microbial contamination. The preparation method is simple and easy, and the required raw materials are relatively easy to obtain. The prepared preservation solution can be stored under appropriate conditions for subsequent use, and its formula is stable and not prone to precipitation or stratification, thereby ensuring the quality and use effect of the preservation solution.

[0022] Preferably, the preservation solution II is prepared by the following preparation method: Weigh 0.024 g of potassium dihydrogen phosphate, 0.014 g of disodium hydrogen phosphate, 0.8 g of sodium chloride, and 0.02 g of potassium chloride and place them in a container, then use 50 ml of ultrapure water to dissolve the solids to obtain.

[0023] By adopting the above technical scheme: the prepared preservation solution II can provide a stable and suitable preservation environment, effectively prolonging the shelf life and use effect of biological molecules, and the raw material cost is relatively low, the preparation process is simple, and the goal of low cost and high benefit is achieved. At the same time, its preparation process is simple and fast, the required raw materials are all common chemicals, and the dosage is precisely controllable, which makes the preparation process easy to standardize and scale, and is convenient for wide application in laboratories or industrial production.

[0024] In summary, the present application includes at least one of the following beneficial technical effects: 1. The kit of the present application comprises magnetic beads coated with different anti-human IgG subtype antibodies (IgG1, IgG2, IgG3, IgG4) and their preservation solution I, as well as acridinium ester-labeled anti-human IgG antibodies and their preservation solution II, which can simultaneously detect four subtypes of IgG antibodies, which provides a more comprehensive and accurate basis for the diagnosis, disease assessment and treatment plan formulation of autoimmune hemolytic anemia (AIHA), and helps doctors to more accurately judge the disease and formulate personalized treatment plans; 2. The kit of this application is used in conjunction with a fully automatic chemiluminescence immunoassay analyzer to achieve automation and standardization of the detection process, which not only reduces human operation errors, improves detection efficiency and repeatability, but also ensures the accuracy and reliability of the test results, which is of great significance for large-scale clinical testing and research; 3. The present application provides a detailed and efficient preparation process from the preparation of magnetic beads coated with anti-human IgG subtype antibodies to the preparation of acridinium ester-labeled anti-human IgG antibodies, and then to the preparation of preservation solution I and preservation solution II. These process steps are clear and easy to operate, suitable for large-scale production and application. At the same time, by precisely controlling the composition and concentration of each reagent, the quality and performance of the kit are ensured, providing strong support for subsequent immunoassays and detection. DETAILED DESCRIPTION

[0025] The present application is further described in detail below in conjunction with the examples. Except for the special instructions below, the raw materials used in the present application are all common commercially available materials. raw material Brand and manufacturer Purpose of Use Specification Anti-human IgG1 subtype antibody Absea Coagulant 1.4mg / mL Anti-human IgG2 subtype antibody Absea Coagulant 1.8 mg / mL Anti-human IgG3 subtype antibody Absea Coagulant 1.5mg / mL Anti-human IgG4 subtype antibody Absea Coagulant 1.2mg / mL Magnetic beads For biological Indicator 3％ Acridinium ester Mei Kate Indicator 2％ Example

[0026] The present application provides a kit for typing and detecting red blood cell IgG antibodies by chemiluminescent immunoassay, the kit comprising reagent R1, reagent R2, reagent R3, reagent R4, and a pre-excitation solution and an excitation solution, wherein reagent R1 is magnetic beads coated with different anti-human IgG subtype antibodies and a preservation solution I thereof, and the concentration of the magnetic beads coated with different anti-human IgG subtype antibodies in the preservation solution I is 0.4 mg / ml; Preservation solution I was prepared by the following preparation method: 1) Use a 1000mL graduated cylinder to measure 800mL of ultrapure water, pour it into a 1000mL beaker, and stir it on a magnetic stirrer; 2) Weigh 3g Tris and 8g NaCl and completely dissolve them in a beaker; 3) Calibrate the pH meter and adjust the pH of the solution in the beaker to 8.0 with 1M HCl solution; 4) Weigh 3 g BSA and dissolve it completely in a beaker; 5) Use a pipette to measure 1 mL of Proclin-300 and add it to the solution and mix well; Pour the solution in the beaker into a 1000mL measuring cylinder, make up to 1000mL, pour into a bottle, and store in a refrigerator at 2-8℃.

[0027] Reagent R2 is an anti-human IgG antibody labeled with acridinium ester and its preservation solution II, and the concentration of the anti-human IgG antibody labeled with acridinium ester in the preservation solution II is 100 ng / ml; Preservation solution II was prepared by the following preparation method: accurately weighed 0.024 g of potassium dihydrogen phosphate, 0.014 g of disodium hydrogen phosphate, 0.8 g of sodium chloride, and 0.02 g of potassium chloride and put them into a 100 ml beaker, 50 ml of ultrapure water was measured with a 100 ml measuring cylinder and added to the beaker to dissolve the solids, the solution was transferred to a 50 ml centrifuge tube, and stored in a refrigerator at 2-8°C.

[0028] Reagents R3 and R4 are both buffer solutions, and are prepared by the following preparation methods: Reagent R3, accurately weigh 3g Tris, 0.8g sodium chloride, 1g BSA, put them into a 100ml beaker, use a 100ml graduated cylinder to measure 50mL ultrapure water and add it to the beaker to dissolve the solid, transfer the solution into a 50ml centrifuge tube, and store in a refrigerator at 2-8℃; Reagent R4, accurately weigh 3g Tris, 0.8g sodium chloride, and 1g casein, and put them into a 100ml beaker. Use a 100ml measuring cylinder to measure 50mL of ultrapure water and add it to the beaker to dissolve the solids. Transfer the solution into a 50ml centrifuge tube and store it in a refrigerator at 2-8℃. The volume of R1 reagent is 50 μl, R2 reagent is 130 μl, R3 reagent is 200 μl, and R4 reagent is 200 μl; Anti-human IgG subtype antibodies include anti-human IgG1 subtype antibodies, anti-human IgG2 subtype antibodies, anti-human IgG3 subtype antibodies, and anti-human IgG4 subtype antibodies. When different anti-human IgG subtype antibodies are coated on magnetic beads, the following steps are taken: 1. Magnetic bead washing: (1) Mix the magnetic beads until they are not sticky to the wall. Take four 2 mL centrifuge tubes and pipette 10 mg of magnetic beads into each tube. Put the others back into the 4 °C refrigerator. (2) Vortex the magnetic beads for 30 seconds, place the centrifuge tube on a magnetic stand to separate the magnetic beads, and remove the supernatant; (3) Add 1 ml of coating buffer, vortex for 30 seconds, clean on an ultrasonic cleaner at 7% ultrasonic power for 1 minute, place on a magnetic stand, wait for 1 minute, and then remove the supernatant of the magnetic beads; (4) Vortex and wash the beads twice with the coating buffer prepared above, and remove the supernatant.

[0029] 2. Magnetic bead coating (1) Add coating buffer to the above centrifuge tube, add 0.2 mg of IgG1, IgG2, IgG3, and IgG4 antibodies respectively, vortex mix, and then add the catalyst and vortex mix evenly; The preparation method of the catalyst is as follows: use a 500mL measuring cylinder to measure 120mL of ultrapure water, pour it into a 500mL beaker, and place it on a magnetic stirrer for stirring; weigh 60g of ammonium sulfate and completely dissolve it in the beaker; calibrate the pH meter and adjust the pH of the solution in the beaker to 9.5 with a 1M NaOH solution; pour the solution in the beaker into a 500mL measuring cylinder, make up to 200mL, pour it into a bottle, and store it in a refrigerator at 2-8°C.

[0030] (2) Incubate the mixture on a rotary mixer at 37°C (20 rpm) for 24 h.

[0031] 3. Magnetic bead blocking (1) Take out the centrifuge tube, adsorb the magnetic beads, and remove the supernatant; (2) Wash the magnetic beads three times with preservation solution I, 1 mL each time; (3) Add 1 mL of Preservation Solution I to the centrifuge tube and incubate at 37°C with rotation (20 rpm).

[0032] 4. Magnetic Bead Cleaning and Storage (1) Take out the magnetic beads, adsorb the magnetic beads, and remove the supernatant; (2) Wash three times with Preservation Buffer I and remove the supernatant.

[0033] (3) Add preservation solution I to the test tube and seal for storage.

[0034] The preparation method of coating buffer is as follows: 1) Use a 500mL graduated cylinder to measure 400mL of ultrapure water, pour it into a 500mL beaker, and stir it on a magnetic stirrer; 2) Weigh 1 g of boric acid and dissolve it completely in a beaker; 3) Calibrate the pH meter and adjust the pH of the solution in the beaker to 8.0 with 1M NaOH solution; 4) Pour the solution in the beaker into a 500mL measuring cylinder and dilute to 500mL; pour into a bottle and store in a refrigerator at 2-8℃.

[0035] Acridinium ester labeled anti-human IgG antibody using the following steps: 1. Take materials: Take 80ul of preservation solution II, 2ul of acridinium ester, and 0.1mg of anti-human IgG antibody and add them into a centrifuge tube, wrap it with tin foil, and place it in a constant temperature and humidity chamber at 37℃ to react for 1h.

[0036] 2. Ultrafiltration to remove free acridinium esters: Use a 50KD ultrafiltration tube to ultrafilter to remove unreacted acridinium ester, add 0.2 mL of preservation solution II, centrifuge at 8000 rpm at 4°C for 5 min, remove the ultrafiltered solution; repeat the above steps twice again, and transfer the labeled antibody to a new centrifuge tube.

[0037] In actual use, cooperate with the fully automatic chemiluminescence immunoassay analyzer to perform sample testing: take out the reagents in the test kit, put them into the reagent drawer, and close the drawer; Place the sample to be tested on the sample rack after opening the cover, and then put the sample rack into the machine; On the test application interface, select "Sample" as the test type, enter the sample information, and select the test item to apply for the test; Click Start to start the test. The reagent dosage for each test is 50μL for R1, 100μL for R2, 100μL for R3, and 100μL for R4. The instrument automatically completes the aspiration and mixing of the sample and reagent R4, and incubates at 37°C. After the incubation is completed, R1 is aspirated and placed into the reaction well, and incubated at 37°C. After the incubation is completed, the immune complex is washed. After washing, R2 and R3 were added and incubated at 37°C. After incubation, the immune complex was washed. After washing, the pre-excitation solution was added and incubated with the immune complex for 1 minute. Then the excitation solution was added and the luminescent signal value was detected. The time from sampling to the result was about 35 minutes.

[0038] Performance testing (I) Physical properties Appearance inspection: Visually inspect the label of the test kit.

[0039] The test results are as follows: The product label of the red blood cell immunoglobulin G subtype detection kit is clearly legible, the outer packaging is not worn, and the text expression is standardized.

[0040] (II) Clinical sample testing The red blood cell immunoglobulin G subtype detection kit prepared in the embodiment is used to detect the red blood cells to be detected, and the detection steps are as follows: (1) adding red blood cells to be tested and magnetic beads coated with anti-human IgG subtype antibodies to the reaction wells, mixing and incubating, the IgG subtype bound to the red blood cells and the anti-human IgG subtype antibodies coated on the magnetic beads bind, washing away the red blood cells that are not bound to the IgG subtype antibodies, and then adding anti-human IgG antibodies labeled with acridinium esters, mixing and incubating, the anti-human IgG antibodies labeled with acridinium esters bind to the IgG antibodies to form an antigen-antibody sandwich complex; (2) After the reaction is completed, the magnetic field attracts the magnetic beads, the cleaning solution washes away the unbound substances, and then the pre-excitation solution and the excitation solution are added to the reaction mixture to cause a chemiluminescent reaction; (3) The number of photons generated in the reaction is measured by a photomultiplier tube, and the number of photons generated is proportional to the concentration of IgG on the red blood cell surface.

[0041] Result determination: The detection limit of this reagent is: 0.1Ru / mL. If the sample concentration value is <0.1Ru / mL, it is judged as negative; if the sample concentration value is ≥0.1Ru / mL, it is judged as positive.

[0042] 1. Reference product L0: physiological saline; Reference product L1: red blood cells of healthy normal people; Reference products L2, L3, L4: red blood cells sensitized by human IgG typing antibodies; Reference product L5: red blood cells sensitized by human IgM antibodies.

[0043] 1.1 Blank limit The reference level L0 was used as the sample to be tested and tested 20 times.

[0044] The test results are as follows: All 10 results were negative (<0.05RU / mL).

[0045] 1.2 Detection limit The reference product L2 was used as the sample to be tested and tested 10 times.

[0046] The test results are as follows: All 10 results were positive (≥0.1RU / mL).

[0047] 1.3 Accuracy The reference product L3 was used as the sample to be tested and tested 10 times.

[0048] The test results are as follows: all 10 results were positive, and the relative deviation of the concentration values ​​did not exceed ±30%.

[0049] The reference product L4 was used as the sample to be tested and tested 10 times.

[0050] The test results are as follows: all 10 results were positive, and the relative deviation of the concentration values ​​did not exceed ±30%.

[0051] 2. Specificity detection The reference product L1 was used as the sample to be tested and tested 10 times; The reference product L2 was used as the sample to be tested and tested 10 times; The reference product L5 was used as the sample to be tested and tested 10 times. The test results are listed in the following table: Table: Specificity test results 3. Repeatability test 3.1 Intra-batch reproducibility The reference product L1 was used as the sample to be tested and tested 10 times; The test results are as follows: 10 results were all negative (<0.1RU / mL) and the coefficient of variation of the concentration value CV ≤ 20.0%; The reference product L3 was used as the sample to be tested and tested 10 times; The test results are as follows: all 10 results were positive (≥0.1RU / mL) and the coefficient of variation of the concentration value CV≤20.0%.

[0052] 3.2 Inter-batch repeatability The intra-batch repeatability was tested using three batches of reagent cards; The test results are consistent with the above results.

[0053] 4. Stability test The product was incubated at 37°C for 7 days before undergoing stability testing to test its physical properties, blank limit, detection limit, accuracy, specificity, and batch-to-batch repeatability; The test results are consistent with the above results.

[0054] In summary, the chemiluminescent immunoassay typing and detection kit for red blood cell IgG antibodies in this application performs well in terms of physical properties, specificity, repeatability and stability, and can meet the clinical needs for IgG antibody subtype detection on red blood cells, providing strong support for the clinical diagnostic accuracy of AIHA.

[0055] The above are all modifications without creative contribution or solutions that obviously constitute technical inspiration that can be made to the present embodiment as needed by technical personnel in this field after reading this specification, but they should be protected by patent law as long as they are within the scope of the claims of this application.

Claims

1. A kit for detecting erythrocyte IgG antibody typing by chemiluminescent immunoassay, It is characterized in that The kit comprises: reagent R1, magnetic beads coated with different anti-human IgG subtype antibodies and preservation solution I thereof; reagent R2, acridinium ester-labeled anti-human IgG antibodies and preservation solution II thereof; reagent R3; reagent R4; pre-excitation solution and excitation solution; wherein the concentration of the magnetic beads coated with different anti-human IgG subtype antibodies in preservation solution I is 0.3-0.4 mg / ml, and the concentration of the acridinium ester-labeled anti-human IgG antibodies in preservation solution II is 95-100 ng / ml.

2. The chemiluminescent immunoassay typing and detection kit for red blood cell IgG antibody according to claim 1, It is characterized in that The volume ratio of the reagent R1, reagent R2, reagent R3 and reagent R4 is 1:(2-2.6):(2-4):(2-4).

3. The chemiluminescent immunoassay typing and detection kit for red blood cell IgG antibody according to claim 1, It is characterized in that The different anti-human IgG subtype antibodies include anti-human IgG1 subtype antibodies, anti-human IgG2 subtype antibodies, anti-human IgG3 subtype antibodies, and anti-human IgG4 subtype antibodies.

4. The chemiluminescent immunoassay typing and detection kit for red blood cell IgG antibody according to claim 1, It is characterized in that The preparation steps of the kit include: a. Preparation of reagent R1: prepare magnetic beads coated with different anti-human IgG subtype antibodies and preservation solution I respectively, then evenly select different types of magnetic beads coated with anti-human IgG subtype antibodies, mix them evenly and put them into preservation solution I, and dilute the anti-IgG subtype antibodies coated with magnetic beads with preservation solution I to 0.4 mg / ml, that is; b. Preparation of reagent R2: prepare acridinium ester-labeled anti-human IgG antibody and preservation solution II respectively, then place the acridinium ester-labeled anti-human IgG antibody into preservation solution II, and dilute the acridinium ester-labeled anti-human IgG antibody to 100 ng / ml with preservation solution II; c. Preparation of reagent R3: Weigh Tris, sodium chloride, and BSA, put them into a beaker, dissolve the solids with ultrapure water, and transfer them to a centrifuge tube to obtain; d. Preparation of reagent R4: Weigh Tris, sodium chloride, and BSA, put them into a beaker, dissolve the solids with ultrapure water, and transfer them to a centrifuge tube.

5. The chemiluminescent immunoassay typing and detection kit for red blood cell IgG antibody according to claim 4, It is characterized in that The method for preparing magnetic beads coated with anti-human IgG subtype antibodies comprises: taking magnetic beads, removing supernatant after vortex separation, washing, adding anti-human IgG antibodies and dispersing them in coating buffer, mixing and incubating; After the incubation is completed, the supernatant is removed, the magnetic beads are washed, and redispersed in the preservation solution I. After blocking and incubation, the result is obtained.

6. The kit for detecting erythrocyte IgG antibody by chemiluminescent immunoassay typing according to claim 4, It is characterized in that The preparation method of the acridinium ester-labeled anti-human IgG antibody is as follows: The anti-human IgG antibody to be labeled and acridinium ester are mixed and dispersed in a labeling buffer, wrapped with tin foil, and allowed to react in a constant temperature and humidity chamber at 35-37°C for 0.5-1h to obtain an anti-human IgG antibody mixture labeled with acridinium ester; Subsequently, the obtained acridinium ester-labeled anti-human IgG antibody mixture is placed in an ultrafiltration tube for ultrafiltration to remove unreacted acridinium ester, thereby obtaining the acridinium ester-labeled anti-human IgG antibody.

7. The kit for detecting erythrocyte IgG antibody by chemiluminescent immunoassay typing according to claim 4, It is characterized in that The preservation solution I is prepared by the following preparation method: Weigh 3g Tris and 8g NaCl into a beaker, and pour in 800ml ultrapure water, stir; adjust the pH of the solution in the beaker to 8.0 with 1M HCl solution; weigh 3g BSA and add it to the solution, and measure 1mL Proclin-300, add it to the solution, mix well and make up to 1000ml.

8. The kit for detecting erythrocyte IgG antibody by chemiluminescent immunoassay typing according to claim 4, It is characterized in that The preservation solution II is prepared by the following preparation method: Weigh 0.024 g of potassium dihydrogen phosphate, 0.014 g of disodium hydrogen phosphate, 0.8 g of sodium chloride, and 0.02 g of potassium chloride and place them in a container, then use 50 ml of ultrapure water to dissolve the solids to obtain.

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