Preparation method of allergen-specific IgE antibody composite quality control product and allergen-specific IgE antibody composite quality control product

An allergen and specific technology, applied in the field of genetic engineering, can solve the problems of easy deterioration or degradation, insufficient supply, cumbersome and difficult adjustment of IgE antibody mixed debugging, and achieve the effect of good stability and high purity

Pending Publication Date: 2021-06-22
湖南携光生物技术有限公司
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

At present, the conventional method of preparing IgE antibody quality control products is mainly to collect allergy-positive serum, which has huge defects. First, positive serum is not easy to collect, the cost is very high, and the supply is often insufficient; The amount is small, and it is difficult to produce large-scale and industrialized products. Third, the stability of natural blood products is poor, and they are easy to deteriorate or degrade during processing, packaging, transportation and

Method used

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  • Preparation method of allergen-specific IgE antibody composite quality control product and allergen-specific IgE antibody composite quality control product
  • Preparation method of allergen-specific IgE antibody composite quality control product and allergen-specific IgE antibody composite quality control product
  • Preparation method of allergen-specific IgE antibody composite quality control product and allergen-specific IgE antibody composite quality control product

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preparation example Construction

[0025] A preparation method of allergen-specific IgE antibody composite quality control product, comprising the following steps:

[0026] S1: Prepare allergen-specific IgE antibodies, as follows: Purify the collected allergen-positive serum with allergen-specific IgE antibody affinity chromatography, elute and collect human IgE; then use the above-mentioned allergen-specific IgE antibodies Affinity chromatography purification of high-purity allergens, elution and collection of allergen-specific IgE antibodies;

[0027] Separation and purification can also be carried out by electrophoresis, salting-out precipitation or other techniques. Total IgE antibodies can be obtained by first purification, and then allergen-specific IgE antibodies can be purified to prevent the mixing of IgG antibodies;

[0028] S2: obtaining the target gene, obtaining the constant region gene sequence and the variable region gene sequence of human IgE;

[0029] The purified allergen-specific IgE antibod...

Embodiment 1

[0048] This example takes the preparation of the composite quality control of shrimp-peanut allergen-specific IgE antibody as an example.

[0049] 1. Preparation of high-purity shrimp-peanut allergen-specific IgE antibody: select one case each of shrimp allergen-positive samples and peanut allergen-positive samples purchased from PlasmaLab in the United States, and mix them with binding buffer (0.02MPBS, pH7.4) volume ratio For 1:1 mixing, 0.8μm membrane filter to remove fibrinogen (to prevent clogging the column). Use 5-10 times the volume of binding buffer to pass through the anti-human IgE antibody column; then load the prepared serum sample, wash the column with binding buffer, and elute the impurity protein; then elute with glycine eluent, and collect at the same time Transudate (about 3-4ml / tube) until there is no protein in the transudate; measure the protein content in each collection tube, combine the protein tubes to obtain the shrimp-peanut allergen-specific IgE ant...

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Abstract

The invention discloses a preparation method of an allergen-specific IgE antibody composite quality control product and the allergen-specific IgE antibody composite quality control product. The preparation method specifically comprises the following steps: preparing an allergen-specific IgE antibody; obtaining a target gene, and obtaining a constant region gene sequence and a variable region gene sequence of human IgE; the method comprises the following steps: designing a primer containing an XhoI restriction enzyme cutting site according to a heavy chain constant region gene sequence of human IgE, designing a primer containing an Hind III restriction enzyme cutting site according to a light chain constant region gene sequence of human IgE, and constructing an expression empty vector containing a light chain signal peptide, a heavy chain signal peptide, a human IgE heavy chain and a light chain C region gene fragment; constructing recombinant expression vectors of different allergens; transfecting the cells; cloning, recognizing and screening; the quality control product has the advantages of high titer, higher purity, favorable dilution linearity and favorable stability, and has higher use value in the aspects of quality control, conventional quality control, performance verification and product development of large-scale production.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to a preparation method of an allergen-specific IgE antibody composite quality control product and the allergen-specific IgE antibody composite quality control product prepared by the method. Background technique [0002] Type I allergic diseases are quite common, and the total incidence rate of the population is as high as 10% to 30%. It is a major worldwide health problem and is listed by the World Health Organization (WHO) as one of the three major diseases that should be prevented and controlled in the 21st century. one. The occurrence of allergic diseases, IgE antibodies play a key role. The detection of allergen-specific IgE antibodies is of great significance in the auxiliary diagnosis, treatment and prevention of allergic diseases. The detection of allergen IgE generally adopts immunodiagnostic methods, such as enzyme-linked immunosorbent assay, flu...

Claims

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Application Information

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IPC IPC(8): C07K16/18C07K16/16C12N15/13C12N15/85G01N33/68G01N33/96
CPCC07K16/18C07K16/16C12N15/85G01N33/6893G01N33/96C07K2317/56G01N2800/24
Inventor 吴荣桂秦枫聂颖王健赵婷季纯宇黄敬双张小飞颜松周玥
Owner 湖南携光生物技术有限公司
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