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Food protein specific IgA antibody detection ELISA kit and application thereof

An antibody detection and specific technology, applied in the biological field, can solve the problems of lack of comparability and unstable human serum standards.

Inactive Publication Date: 2018-12-18
SOOCHOW UNIV AFFILIATED CHILDRENS HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no ELISA kit for the detection of food-specific IgA antibodies on the market
Some of the ELISA systems for detecting food-specific IgA antibodies in other diseases reported in previous literatures have no standard products, and only compare the absorbance of the samples to be tested, and the absorbance of different ELISA plates is not comparable; some studies Use human serum or plasma to establish standards, or use a subject’s serum as a reference to standardize the absorbance values ​​of samples between different microtiter plates, while human serum is unstable as a standard

Method used

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  • Food protein specific IgA antibody detection ELISA kit and application thereof
  • Food protein specific IgA antibody detection ELISA kit and application thereof
  • Food protein specific IgA antibody detection ELISA kit and application thereof

Examples

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preparation example Construction

[0059] The preparation method of the antigen-coated microtiter plate disclosed by the invention comprises the following steps:

[0060] (1) Coating: Dilute the antigen to an appropriate concentration with pH 9.6 carbonate buffer solution, 50 μl / well, cover the microtiter plate with parafilm, and place it at 4°C overnight; when the antigen is bovine β-lactoglobulin or bovine serum albumin When one of protein and ovalbumin is used, the antigen concentration is 1 μg / mL; when the antigen is bovine casein, the antigen concentration is 0.1 μg / mL;

[0061] (2) Plate washing: Pour off the coating solution, wash the plate 3 times with PBST, each time gently shake off the washing solution on the sink, and pat the microplate on a paper towel to remove the residual liquid;

[0062] (3) Sealing: Add protein-free blocking solution to each well to seal the space on the microplate plate that is not adsorbed by the target protein, 200 μl / well, cover the microplate plate with a sealing film, an...

Embodiment 1

[0078] Determination of the dilution of biotin antibody, the dilution of biotin secondary antibody in the ELISA kit system determined in this example is: casein: 1:100,000, β-lactoglobulin: 1:200,000, bovine serum albumin : 1:10,000, ovalbumin: 1:200,000.

[0079] Determination of the dilution of the sample to be tested, the dilution of the sample to be tested in the ELISA kit system determined in this embodiment is respectively: casein: 1:400, β-lactoglobulin: 1:400, bovine serum albumin: 1: 1600, ovalbumin: 1:400.

[0080] The standard substance concentration gradient determined in this example is: casein and bovine serum albumin-specific IgA antibodies corresponding to the human total IgA antibody dilution: 200 μg / ml, 100 μg / ml, 50 μg / ml, 25 μg / ml, 12.5 μg / ml ml, 6.25 μg / ml, 3.125 μg / ml, 0 μg / ml. The dilutions of human total IgA antibodies corresponding to β-lactoglobulin and ovalbumin-specific IgA antibodies are: 400 μg / ml, 200 μg / ml, 100 μg / ml, 50 μg / ml, 25 μg / ml, 12.5 ...

Embodiment 2

[0096] Embodiment 2 The minimum detection limit (sensitivity) of the kit of the present invention

[0097] Determination of the dilution of biotin antibody, the dilution of biotin secondary antibody in the ELISA kit system determined in this example is: casein: 1:100,000, β-lactoglobulin: 1:200,000, bovine serum albumin : 1:10,000, ovalbumin: 1:200,000.

[0098] Determination of the dilution of the sample to be tested, the dilution of the sample to be tested in the ELISA kit system determined in this embodiment is respectively: casein: 1:400, β-lactoglobulin: 1:400, bovine serum albumin: 1: 1600, ovalbumin: 1:400.

[0099] The standard substance concentration gradient determined in this example is: casein and bovine serum albumin-specific IgA antibodies corresponding to the human total IgA antibody dilution: 200 μg / ml, 100 μg / ml, 50 μg / ml, 25 μg / ml, 12.5 μg / ml ml, 6.25 μg / ml, 3.125 μg / ml, 0 μg / ml. The dilutions of human total IgA antibodies corresponding to β-lactoglobulin ...

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Abstract

The invention discloses a food protein specific IgA antibody detection ELISA kit and an application thereof. The food protein specific IgA antibody detection ELISA kit comprises diluent, an antigen-coated ELISA plate, a plate washing solution, a standard product, negative control, a biotin secondary antibody, an avidin-HRP solution, a developing solution and stop buffer . The kit can be applied tothe detection of patients with IgA-related diseases such as allergic purpura (IgA vasculitis), IgA nephropathy, rheumatoid arthritis, juvenile idiopathic arthritis, systemic lupus erythematosus and the like so as to explore the correlation between the patients and food allergy proteins, which is conducive to the prevention and treatment of diseases. At the same time, the food protein specific IgAantibody detection ELISA kit satisfies the quality control standards of clinical food specific antibody detection kits (enzyme-linked immunosorbent assay) used in domestic hospitals, as well as the ELISA principle, characteristics and influence factors.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to an ELISA kit for detecting food protein-specific IgA antibodies and an application thereof. Background technique [0002] At present, ELISA kits for food-specific IgE and IgG antibodies have been widely used in clinical practice. However, there is no ELISA kit for detecting food-specific IgA antibodies on the market. Some of the ELISA systems for detecting food-specific IgA antibodies in other diseases reported in previous literatures have no standard products, and only compare the absorbance of the samples to be tested, and the absorbance of different ELISA plates is not comparable; some studies Use human serum or plasma to establish a standard, or use a subject's serum as a reference to standardize the absorbance values ​​of samples between different microtiter plates, and human serum is unstable as a standard. Contents of the invention [0003] The present inventio...

Claims

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Application Information

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IPC IPC(8): G01N33/68
CPCG01N33/68G01N33/6854
Inventor 李晓忠王凤英缪美华胡筱涵林强朱赟陈如月
Owner SOOCHOW UNIV AFFILIATED CHILDRENS HOSPITAL
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