IL-13 detection kit based on magnetic particle chemiluminescence method, preparation method and application

Abstract

The invention provides an IL-13 detection kit based on a magnetic particle chemiluminescence method, and belongs to the technical field of medical detection.The kit comprises a reagent R1, a reagent R2, a reagent R3 and a reagent R4, the reagent R1 comprises a magnetic particle suspension, the reagent R2 comprises an analysis buffer solution, the reagent R3 comprises an IL-13 antibody solution, and the reagent R4 comprises a magnetic particle solution. The IL-13 antibody solution contains a biotin-labeled IL-13 antibody, the concentration of the biotin-labeled IL-13 antibody is 0.1-1.0 mg / mL, and the labeling ratio of the IL-13 antibody to biotin is 1: (5-50); and the reagent R4 comprises a streptavidin buffer solution. The kit is adaptive to a chemiluminescence instrument, can rapidly and effectively detect the concentration of IL-13, and improves the sensitivity, accuracy and detection efficiency of IL-13 detection. The invention further provides a preparation method and application of the IL-13 detection kit based on the magnetic particle chemiluminescence method.

Description

Technical Field

[0001] The invention belongs to the technical field of medical detection, and particularly relates to an IL-13 detection kit based on magnetic particle chemiluminescence method, a preparation method and application thereof. Background Art

[0002] Interleukin 13 was discovered in 1989 and is known as the P600 protein. Like all other cytokines, IL-13 is a pleiotropic cytokine. IL-13 acts on normal and malignant B cells, monocytes, macrophages, NK cells, polymorphonuclear cells, osteoblasts, endothelial cells, fibroblasts, and keratinocytes, indicating that it not only has important immunomodulatory properties, but also has the function of regulating a variety of other cell types.

[0003] The cDNA of IL-13 encodes a protein of 132 amino acids with four potential N-linked glycosylation sites and five cysteine ​​residues, four of which can form disulfide pairs. IL-13 is mainly produced by CD4+ cells and secreted by several helper T cell subsets, NK cells, mast cells, eosinophils, basophils, and visceral smooth muscle cells. It plays a key role in the pathogenesis of allergies, asthma, cancer, and tissue fibrosis.

[0004] For the detection of IL-13, there is currently no kit using the magnetic microparticle luminescent immunoassay method. At the same time, the detection time is greatly shortened compared to the enzyme immunoassay (the enzyme immunoassay generally takes 4-5 hours to produce results, and the use of the magnetic microparticle luminescent immunoassay method is expected to shorten the detection time to 30 minutes). In addition, the abundance of IL-13 in serum is low (fg / ml level) and is not easy to be detected, and it is difficult to detect accurately and repeatedly. Summary of the invention

[0005] In order to solve the problems of low sensitivity and low detection efficiency of existing IL-13 detection kits, the present invention provides an IL-13 detection kit based on magnetic particle chemiluminescence method, which is adapted to a chemiluminometer and can quickly and effectively detect IL-13 concentration, thereby improving the sensitivity, accuracy and detection efficiency of IL-13 detection.

[0006] The present invention also provides a preparation method and application of an IL-13 detection kit based on the magnetic particle chemiluminescence method.

[0007] The present invention is achieved through the following technical solutions:

[0008] The present invention provides an IL-13 detection kit based on magnetic particle chemiluminescence method, the kit comprising reagent R1, reagent R2, reagent R3 and reagent R4;

[0009] The reagent R1 includes a magnetic particle suspension, and the magnetic particle suspension contains the following components:

[0010] Magnetic particles with IL-13 antibody coupled to the surface 0.1-1.0 mg / ml; Tris 40-60 mM; Nacl 90-120 mM; bovine serum albumin 1-2 wt%; glycerol 4-7 wt%; sucrose 2-5 wt%; Proclin300 0.05-0.2 wt%;

[0011] The reagent R2 includes an analysis buffer, which contains the following components:

[0012] BIS-TRIS 40~60mM; Triton X-100 0.1~0.2wt%; Polyethylene glycol-6000 0.03~0.05wt%; Dextran sulfate sodium 0.05~0.1wt%; Hexadecyltrimethylammonium chloride 0.1~0.2wt%; Proclin 300 0.1~0.2wt%; Mouse IgG 0.1~0.2mg / mL;

[0013] The reagent R3 includes an IL-13 antibody solution, wherein the IL-13 antibody solution contains a biotin-labeled IL-13 antibody, wherein the concentration of the biotin-labeled IL-13 antibody is 0.1 to 1.0 mg / mL, and the labeling ratio of the IL-13 antibody to biotin is 1:5 to 50;

[0014] The reagent R4 includes a streptavidin buffer, and the streptavidin buffer contains the following components:

[0015] Acridinium ester-labeled streptavidin 0.1±0.01μg / mL; Nacl 0.1~0.15M; Proclin300 0.1~0.2wt%; Triton X-100 0.1~0.3wt%; Phosphate 10~20mM;

[0016] In the acridinium ester-labeled streptavidin, the labeling ratio of acridinium ester to streptavidin is 1:10-100. (i.e., the molar ratio of acridinium ester to streptavidin is 1:10-100)

[0017] Furthermore, the IL-13 antibody solution contains the following components:

[0018] Biotin-labeled IL-13 antibody 0.1-1.0 mg / mL; PB 0.01-0.02 wt%; BSA 1±0.1 wt%; Proclin 300 0.1±0.01 wt%; protective protein ADP 0.1±0.01 wt%; NP 40 0.1±0.01 wt%; Dinazol 20±1 ng / mL.

[0019] Furthermore, the kit also includes a calibrator, which is obtained by diluting the IL-13 antigen with a calibrator diluent.

[0020] Furthermore, the calibration diluent comprises a phosphate buffer, and the phosphate buffer contains the following components:

[0021] Protective protein ADP 0.1wt%; BSA (bovine serum albumin) 2wt%; phosphate 20mmol / L;

[0022] The IL-13 antigen concentration gradient of the calibrator includes 0pg / mL, 5pg / mL, 10pg / mL, 20pg / mL, 50pg / mL and 100pg / mL.

[0023] Furthermore, the particle size of the magnetic particles with IL-13 antibody coupled to the surface is 0.1 to 3.0 μm;

[0024] The magnetic particles include hydrophilic magnetic beads MS160.

[0025] Based on the same inventive concept, the present invention provides a method for preparing an IL-13 detection kit based on magnetic particle chemiluminescence, the preparation method comprising:

[0026] Preparation of reagent R1:

[0027] The activated magnetic particles are coupled with IL-13 antibodies to obtain magnetic particles with IL-13 antibodies coupled to the surface;

[0028] The magnetic particles with IL-13 antibody coupled to the surface are mixed with Tris, NaCl, bovine serum albumin, glycerol, sucrose, and Proclin 300 solution to obtain reagent R1;

[0029] Preparation of reagent R2:

[0030] Prepare 40-60 mM BIS-TRIS (bis(2-hydroxymethyl)amino-tris(hydroxymethyl)methane) buffer, add Triton X-100, polyethylene glycol-6000, dextran sodium sulfate, hexadecyltrimethylammonium chloride, Proclin 300 solution and mouse IgG, mix well, and obtain reagent R2;

[0031] Preparation of reagent R3:

[0032] The IL-13 antibody dialyzed with PBS buffer and long-chain biotin were mixed at a molar ratio of 1:5-50, and the labeled substance was collected after dialysis to obtain reagent R3.

[0033] Preparation of reagent R4:

[0034] Acridinium ester and streptavidin are mixed and reacted in the dark to obtain streptavidin labeled with acridinium ester;

[0035] The acridinium ester-labeled streptavidin is mixed with NaCl, Proclin 300 solution, and a phosphate buffer containing Triton X-100 to obtain reagent R4;

[0036] Preparation of calibrators:

[0037] Using 20 mM phosphate buffer containing 0.1% protective protein ADP and 2% BSA, solutions with IL-13 antigen concentrations of 0 pg / mL, 5 pg / mL, 10 pg / mL, 20 pg / mL, 50 pg / mL and 100 pg / mL were prepared to obtain calibrants.

[0038] Based on the same inventive concept, the present invention provides an application of an IL-13 detection kit based on magnetic particle chemiluminescence method in in vitro detection of IL-13.

[0039] One or more technical solutions in the embodiments of the present invention have at least the following technical effects or advantages:

[0040] 1. The present invention discloses an IL-13 detection kit based on magnetic particle chemiluminescence method, which adopts biotin-avidin system and increases analysis buffer component, wherein hexadecyltrimethylammonium chloride, a substance capable of enhancing reactivity and signal value, is added, which can eliminate interference in the reaction, reduce matrix difference and enhance signal value level, thereby improving the sensitivity of reagent detection, and can be traced back to international standard products, thus ensuring product accuracy and high consistency, which is conducive to mutual recognition of results of different laboratories and manufacturers, and greatly shortens the detection time compared with enzyme immunoassay detection time.

[0041] 2. The present invention provides an IL-13 detection kit based on magnetic microparticle chemiluminescence method, which can be adapted to a fully automatic chemiluminescence instrument. It has the advantages of simple operation, high sensitivity, low cost, short operation time, stable results, and can effectively improve the detection efficiency and accuracy, providing reference opinions for clinical doctors in clinical diagnosis. BRIEF DESCRIPTION OF THE DRAWINGS

[0042] In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the following briefly introduces the drawings required for use in the description of the embodiments. Obviously, the drawings described below are some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without creative work.

[0043] Figure 1 This is a diagram of the structure of IL-13 markers and its receptor protein.

[0044] Figure 2 The figure is the correlation between the kit of the present invention and the ELISA kit. DETAILED DESCRIPTION

[0045] The present invention will be described in detail below in conjunction with specific implementations and examples, and the advantages and various effects of the present invention will be more clearly presented. It should be understood by those skilled in the art that these specific implementations and examples are used to illustrate the present invention, rather than to limit the present invention.

[0046] Throughout the specification, unless otherwise specifically stated, the terms used herein should be understood as meanings commonly used in the art. Therefore, unless otherwise defined, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art to which the present invention belongs. In the event of a conflict, the present specification takes precedence.

[0047] Unless otherwise specified, various raw materials, reagents, instruments and equipment used in the present invention can be purchased from the market or prepared by existing methods.

[0048] The following is a detailed description of an IL-13 detection kit based on magnetic microparticle chemiluminescence method of the present application in combination with examples and experimental data.

[0049] Example 1

[0050] This embodiment provides a method for preparing an IL-13 detection kit based on magnetic microparticle chemiluminescence method, which is as follows:

[0051] 1. Preparation of reagent R1:

[0052] The magnetic microparticles (hydrophilic magnetic beads MS160, purchased from JSR) (particle size 0.16 μm-0.3 μm) were washed 3 times with MES buffer, activated for 30-60 min, washed twice with MES buffer, and then IL-13 antibody (mouse monoclonal antibody, purchased from Baixinyi Biotechnology Co., Ltd.) was added for coupling to couple the antibody to the surface of the magnetic microparticles. Then, 50 mM Tris buffer (containing 0.05% Tween 20) was used as a washing buffer, and the magnetic microparticles were washed twice, and then a blocking solution was added for blocking. The magnetic microparticles were washed twice with 50 mM Tris buffer (containing 0.05% Tween 20), and then a blocking solution containing 50 mM Tris, 100 mM A buffer solution containing NaCl, 1% bovine serum albumin, 5% glycerol, 2% sucrose and 0.1% Proclin300 was prepared into a magnetic particle working solution with a magnetic particle suspension concentration (i.e., magnetic particle concentration) of 0.5 mg / ml, which was sealed in a reagent bottle to obtain R1 reagent.

[0053] 2. Preparation of reagent R2:

[0054] Prepare 50mM BIS-TRIS buffer, add 0.1% Triton X-100, 0.05% polyethylene glycol-6000 (PEG6000), 0.1% dextran sulfate sodium, 0.2% hexadecyltrimethylammonium chloride (CTAC), and then add 0.1% Proclin300 and mouse IgG0.2mg / ml (all final concentrations or final mass fractions), mix thoroughly, and fill in a reagent bottle to obtain R2 reagent.

[0055] 3. Preparation of reagent R3:

[0056] The IL-13 antibody was dialyzed in PBS buffer overnight, and the dialyzed IL-13 antibody and long-chain biotin (purchased from Solebol) were mixed at a molar ratio of 1:10 and dialyzed overnight. After overnight, the marker was collected and diluted with a buffer (PBS+BSA+Proclin300+ADP) to prepare reagent R3.

[0057] 4. Preparation of reagent R4:

[0058] Add streptavidin to carbonate buffer solution and mix thoroughly. After mixing, add 2.0 mg / ml acridinium ester solution dissolved in anhydrous DMSO, vortex mix for 30 seconds, seal and mix in the dark for 2 hours; add 25% lysine blocking solution, mix and block for 2 hours; purify and collect the blocked labeled streptavidin by ammonium sulfate precipitation method, and dilute the obtained acridinium ester-labeled streptavidin with 20 mM phosphate buffer containing 100 mM Nacl, 0.1% Proclin300, and 0.1% Triton X-100 to a final concentration of 0.1 μg / mL, and fill it in a reagent bottle to obtain R4 reagent.

[0059] 5. Preparation of Calibrators:

[0060] Use 20 mM phosphate buffer containing 0.1% protective protein ADP, 0.1% Proclin300, and 2% BSA to prepare IL-13 calibrators with concentrations of 0 pg / mL, 5 pg / mL, 10 pg / mL, 20 pg / mL, 50 pg / mL, and 100 pg / mL.

[0061] Example 2

[0062] This example is used to screen the components of the kit.

[0063] 1. Screening of reagent R2:

[0064] 1.1 Comparison of different buffers;

[0065] Buffers include PBS buffer (pH 7.2), Tris buffer (pH 7.4), Bis-Tris buffer (pH 7.4), and MES buffer (pH 6.5);

[0066] Table 1: Test results of R2 in different buffers

[0067] PBS buffer Tris Buffer Bis-Tris Buffer MES Buffer 0 value 1245 954 758 1354 S1 7854 8214 9874 7598 S2 14587 15846 16785 13584 S3 31024 33254 32580 30254 S4 79854 81574 83728 75647 S5 168745 17548 180761 154785

[0068] Conclusion: Comparing different buffers, it can be seen from the results that Bis-Tris buffer has a slightly higher signal value level than other buffers, and the 0 value is lower, so this buffer was selected (different buffer selections for R2, calibration products S1-S5 and background 0 value detection and comparison under a certain buffer, the background 0 value and calibration products S1-S5 components are: 20mMPB+2%BSA+0.1%ADP+0.1%Proclin300, IL-13 concentrations are 0pg / mL, 5pg / mL, 10pg / ml, 20pg / ml, 50pg / mL, 100pg / mL respectively).

[0069] 1.2 Reagent R2 formula screening

[0070] Formula 1Bis-Tris+BSA+PEG-6000+Triton+Proclin300+mouse IgG;

[0071] Formula 2: Bis-Tris+BSA+PEG-6000+Triton+Proclin300+mouse IgG+0.1%DSS;

[0072] Formula 3: Bis-Tris+BSA+PEG-6000+Triton+Proclin300+mouse IgG+0.05% CTAC;

[0073] Formula 4: Bis-Tris+BSA+PEG-6000+Triton+Proclin300+mouse IgG+0.1%DSS+0.05%CTAC;

[0074] The reagents were brought into the instrument for testing. The instrument was KunLun i2000. The test results are shown in Table 2:

[0075] Table 2: R2 component formulation screening

[0076] sample Recipe 1 Recipe 2 Recipe 3 Recipe 4 0 value 345 457 384 485 S1 8524 10824 13422 24159 S2 16542 19520 24204 42357 S3 33245 38232 48937 82214 S4 85437 97996 121515 167691 S5 182587 212349 256942 472773 P1 4774 6312 7572 13777 P2 1838 2618 2919 5468 P3 3986 5352 6350 11650

[0077] From the results, it can be seen that adding DSS and CTAC to the components has a significant effect on improving the signal value and thus improving the sensitivity (P1-P3 are clinical samples), and the mixed use of the two can significantly increase the signal value.

[0078] 1.3; Comparison of adding DSS and CTAC in different proportions:

[0079] Formula 1Bis-Tris+1%BSA+0.1%Proclin300+0.1%Triton

[0080] X-100+0.05%PEG-6000+0.05%DSS+0.05%CTAC;

[0081] Formula 2Bis-Tris+1%BSA+0.1%Proclin300+0.1%Triton X-100+0.05%PEG-6000+0.1%DSS+0.05%CTAC;

[0082] Formula 3Bis-Tris+1%BSA+0.1%Proclin300+0.1%Triton X-100+0.05%PEG-6000+0.1%DSS+0.1%CTAC;

[0083] Table 3: DSS and CTAC at different ratios

[0084] Recipe 1 Recipe 2 Recipe 3 0 value 425 397 541 S1 22478 28474 28947 S2 40784 46587 47851 S3 78547 83547 84678 S4 145874 175844 185475 S5 385475 489751 479612

[0085] From the results, it can be seen that when 0.1% DSS and 0.05% CTAC are added, the signal value is basically at a plateau, so both can be added according to this amount.

[0086] 1.4 Reagent R3 component buffer

[0087] Nonylphenol polyoxyethylene ether NP40 surfactant is added to the R3 component, and Dinazol (danazol) dissociates the marker binding receptor, thereby improving the sensitivity of the reagent for marker detection;

[0088] The comparison data before and after adding is as follows:

[0089] Formula before addition: PBS+1%BSA+0.1%Proclin300+0.1%ADP;

[0090] Formula after addition: PBS + 1% BSA + 0.1% Proclin300 + 0.1% ADP + 0.1% nonylphenol polyoxyethylene ether NP40 surfactant + 20ng / mL Dinazol;

[0091] The test results are shown in Table 4.

[0092] Table 4: R3 component formulation comparison

[0093] sample Before adding After adding 0 value 425 510 S1 27050 25698 S2 45189 43834 S3 81876 80239 S4 167052 158699 S5 499546 494551 Serum 1 1235 1587 Serum 2 5748 8622 Serum 3 12698 17142 Serum 4 9547 12220

[0094] It can be concluded from Table 4 that NP40 and Dinazol can be added to the R3 component to improve the detection sensitivity.

[0095] Example 3

[0096] This example performs a performance test on the kit of the present invention, as follows:

[0097] 1. Sensitivity test

[0098] Limit of Blank (LOB): 5 blank samples were tested 3 times per day for 4 consecutive days, and a total of 60 data points were obtained.

[0099] Limit of detection (LOD): After the LOB was determined, five clinical samples with low values ​​of 1-4 times the LOB were tested three times for each sample and tested continuously for 4 days, resulting in a total of 60 data points.

[0100] Limit of quantitation (LOQ): Based on the 12 test results of each low-concentration sample of each batch of reagents when the upper L0D is established, calculate the mean, standard deviation and bias of the sample, and calculate TE (%) according to the formula TE (%) = (|bias| + 2*standard deviation) / reference value × 100%; compare the TE of each sample with the TE target value, and select the lowest concentration that meets the TE target value as the estimated value of the LoQ of the batch.

[0101] The test results are shown in Table 5:

[0102] Table 5 Sensitivity test results of this kit

[0103]

[0104] As can be seen from Table 5, the sensitivity of the kit of the present invention can reach 0.04 pg / ml. (Batch 1 and batch 2 in the table refer to two batches of reagents prepared after the final formula is determined, and there is no batch difference between the two batches of reagents)

[0105] 2. Kit precision test

[0106] Two batches of reagents were taken for precision testing, using high and low quality control and clinical low, medium and high samples respectively. The measurement was repeated 20 times, and the variation of the measured concentration was calculated. The measurement results are shown in Tables 6 and 7.

[0107] Table 6 Quality control product precision test results

[0108]

[0109]

[0110] Table 7 Clinical precision test

[0111]

[0112]

[0113] The results in Table 6 and Table 7 show that the coefficient of variation CV is less than 8.0%. (Sample concentration 1-20 means 20 replicates)

[0114] 3. Kit cross-reaction assessment

[0115] The interference of interleukin-2, interleukin-4, interleukin-8, interleukin-5, interleukin-2R, interleukin-6, and interleukin-17A on the kit of the present invention was assessed. A diluent with a ratio of no more than 5% was added as a control sample, and a ratio of no more than 5% of interleukin 2 (IL-2) antigen pure product, interleukin 4 (IL-4) antigen pure product, interleukin 8 (IL-8) antigen pure product, interleukin 5 (IL-5) antigen pure product, interleukin 2 receptor (IL-2R) antigen pure product, interleukin 6 (IL-6) antigen pure product, and interleukin 17A (IL-17A) antigen pure product were added, and the mixture was prepared to contain 10000.0 pg / mL IL- 2, cross-product 2 with 10000.0pg / mL IL-4, cross-product 3 with 10000.0pg / mL IL-8, cross-product 4 with 10000.0pg / mL IL-5, cross-product 5 with 10000.0pg / mL IL-2R, cross-product 6 with 10000.0pg / mL IL-6, and cross-product 7 with 10000.0pg / mL IL-17A were used as cross-samples, and the cross-reaction rate was calculated according to the following formula.

[0116]

[0117] Measurement value – the measurement mean of the added sample;

[0118] Theoretical value – the mean value of the control sample;

[0119] Interferent Concentration – The concentration of the added interferent.

[0120] The results are shown in Table 8 below:

[0121] Table 8 Cross-reaction

[0122]

[0123] The results in Table 8 show that the crossover rates between the kit of the present invention and the main crossover substances are all less than 0.1%, and there is no crossover to the kit of the present invention.

[0124] 4. Clinical performance assessment of the test kit

[0125] The kit of the present invention and the scientific research ELISA kit were used to simultaneously test samples from 20 patients with clinical diagnosis of allergic reactions, asthma and other inflammations (the 20 samples were divided into two parts, one was tested with the kit of the present invention, and the other was tested with the ELISA kit according to its operating instructions). The test correlation is shown in Figure 2 ,according to Figure 2 The correlation results show that the correlation R2 between the two is 0.9884, which is a good correlation.

[0126] The kit of the present invention adopts a biotin-avidin system, where one avidin can bind to four biotins, which can improve sensitivity compared to direct coating (one antigen determinant binds to one antibody); hexadecyltrimethylammonium chloride is a surfactant, and can also enhance the luminescence of acridinium ester; the addition of NP40 surfactant and Dinazol may play a role in the separation of the receptor bound to the marker ( Figure 1 ).

[0127] Finally, it should be noted that the terms "comprises," "includes," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that includes a series of elements includes not only those elements, but also other elements not explicitly listed, or also includes elements that are inherent to such process, method, article, or apparatus.

[0128] Although the preferred embodiments of the present invention have been described, those skilled in the art may make other changes and modifications to these embodiments once they have learned the basic creative concept. Therefore, the appended claims are intended to be interpreted as including the preferred embodiments and all changes and modifications that fall within the scope of the present invention.

[0129] Obviously, those skilled in the art can make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if these modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include these modifications and variations.

Claims

1. An IL-13 detection kit based on magnetic microparticle chemiluminescence method, characterized in that: The kit comprises reagent R1, reagent R2, reagent R3 and reagent R4; The reagent R1 includes a magnetic particle suspension, and the magnetic particle suspension contains the following components: Magnetic particles with IL-13 antibody coupled to the surface 0.1-1.0 mg / ml; Tris 40-60 mM; Nacl 90-120 mM; bovine serum albumin 1-2 wt%; glycerol 4-7 wt%; sucrose 2-5 wt%; Procl in300 0.05-0.2 wt%; The reagent R2 includes an analysis buffer, which contains the following components: BIS-TRIS 40~60mM; Triton X-100 0.1~0.2wt%; Polyethylene glycol-6000 0.03~0.05wt%; Dextran sulfate sodium 0.05~0.1wt%; Hexadecyltrimethylammonium chloride 0.1~0.2wt%; Procl in300 0.1~0.2wt%; Mouse IgG 0.1~0.2mg / mL; The reagent R3 includes an IL-13 antibody solution, wherein the IL-13 antibody solution contains a biotin-labeled IL-13 antibody, wherein the concentration of the biotin-labeled IL-13 antibody is 0.1 to 1.0 mg / mL, and the labeling ratio of the IL-13 antibody to biotin is 1:5 to 50; The reagent R4 includes a streptavidin buffer, and the streptavidin buffer contains the following components: Acridinium ester labeled streptavidin 0.1±0.01μg / mL; NaCl 0.1~0.15M; Procl in300 0.1~0.2wt%; Triton X-100 0.1~0.3wt%; Phosphate 10~20mM; In the acridinium ester-labeled streptavidin, the labeling ratio of acridinium ester to streptavidin is 1:10-100.

2. The IL-13 detection kit based on magnetic microparticle chemiluminescence method according to claim 1, characterized in that: The IL-13 antibody solution contains the following ingredients: Biotin-labeled IL-13 antibody 0.1-1.0 mg / mL; PB 0.01-0.02 wt%; BSA 1±0.1 wt%; Proclin 300 0.1 ± 0.01 wt%; Protective protein ADP 0.1 ± 0.01 wt%; NP400.1±0.01wt%; Dinazol20±1ng / mL.

3. The IL-13 detection kit based on magnetic microparticle chemiluminescence method according to claim 1, characterized in that: The kit also includes a calibrator, which is obtained by diluting the IL-13 antigen with a calibrator diluent.

4. The IL-13 detection kit based on magnetic microparticle chemiluminescence method according to claim 3, characterized in that: The calibration diluent comprises a phosphate buffer, wherein the phosphate buffer contains the following components: Protective protein ADP 0.1wt%; BSA 2wt%; Phosphate 20mmol / L; The IL-13 antigen concentration gradient of the calibrator includes 0pg / mL, 5pg / mL, 10pg / mL, 20pg / mL, 50pg / mL and 100pg / mL.

5. The IL-13 detection kit based on magnetic microparticle chemiluminescence method according to claim 1, characterized in that: The particle size of the magnetic particles with IL-13 antibody coupled to the surface is 0.1 to 3.0 μm; The magnetic particles include hydrophilic magnetic beads MS160.

6. A method for preparing an IL-13 detection kit based on magnetic particle chemiluminescence method according to any one of claims 1 to 5, characterized in that: The preparation method comprises: Preparation of reagent R1: The activated magnetic particles are coupled with IL-13 antibodies to obtain magnetic particles with IL-13 antibodies coupled to the surface; The magnetic particles with IL-13 antibody coupled to the surface are mixed with Tris, NaCl, bovine serum albumin, glycerol, sucrose, and Proclin 300 solution to obtain reagent R1; Preparation of reagent R2: Prepare 40-60 mM BIS-TRIS buffer, add Triton X-100, polyethylene glycol-6000, dextran sodium sulfate, hexadecyltrimethylammonium chloride, Proclin 300 solution and mouse IgG, mix well to obtain reagent R2; Preparation of reagent R3: The IL-13 antibody dialyzed with PBS buffer was mixed with long-chain biotin at a molar ratio of 1:5-50, and the labeled substance was collected after dialysis to obtain reagent R3. Preparation of reagent R4: Acridinium ester and streptavidin are mixed and reacted in the dark to obtain streptavidin labeled with acridinium ester; The acridinium ester-labeled streptavidin is mixed with NaCl, Proclin 300 solution, and a phosphate buffer containing Triton X-100 to obtain reagent R4; Preparation of calibrators: Using 20 mM phosphate buffer containing 0.1% protective protein ADP and 2% BSA, solutions with IL-13 antigen concentrations of 0 pg / mL, 5 pg / mL, 10 pg / mL, 20 pg / mL, 50 pg / mL and 100 pg / mL were prepared to obtain calibrants.

7. Use of an IL-13 detection kit based on magnetic microparticle chemiluminescence method according to any one of claims 1 to 5 in detecting IL-13 in vitro.

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