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A kind of rabies virus recombinant antigen and preparation method and use thereof

A rabies virus, recombinant antigen technology, applied in the directions of virus antigen components, chemical instruments and methods, biochemical equipment and methods, etc., can solve the problems of low sensitivity, high preparation cost, large infection risk, etc. The effect of good antigenicity and reducing the risk of anti-virus

Active Publication Date: 2022-07-12
北京世纪元亨动物防疫技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the whole virus antigen detection kit has the advantage of high specificity in detecting rabies antibodies, there is a great risk of infection for the staff during the preparation and later purification of the whole virus antigen, and the preparation cost is high and the amount obtained is small
However, when using nucleoprotein antigen or glycoprotein antigen detection kits alone to detect rabies antibodies, there are deficiencies such as missed detection, poor specificity, and low sensitivity of the method.

Method used

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  • A kind of rabies virus recombinant antigen and preparation method and use thereof
  • A kind of rabies virus recombinant antigen and preparation method and use thereof
  • A kind of rabies virus recombinant antigen and preparation method and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0085] Example 1 Preparation of rabies virus recombinant antigen

[0086] (1) Primer design

[0087] Use DNAman and DNAstar to analyze the gene of amino acid sequence fragment N (as shown in SEQ ID NO. 4), the gene of amino acid sequence fragment G (as shown in SEQ ID NO. 5) and the gene of amino acid sequence fragment P (as shown in SEQ ID NO. 6), 3 pairs of primers (primer 1F and primer 1R, primer 2F and primer 2R, primer 3F and primer 3R) were designed to amplify the genes of amino acid sequence fragments N, P and G, respectively. In order to facilitate the subsequent concatenation of the amplified three gene fragments, and to facilitate each gene to form its own specific epitope after concatenation, the nucleotide sequence of the flexible peptide and the amino acid of the flexible peptide are added to the primers between the gene linkages. The sequence is GGGSSS, GGSGGS or GGGGS. In this example, the flexible peptide GGGGS is selected. For details, see the double underlin...

Embodiment 2

[0120] Embodiment 2 A kind of rabies antibody detection test strip and preparation method thereof

[0121] This embodiment provides a rabies antibody detection test strip, such as Image 6 As shown, it includes: a bottom plate 7 and a sample pad 1, a gold standard pad 2, a nitrocellulose membrane 3 and a water-absorbing pad 6 that are sequentially adhered to the bottom plate along the length direction of the bottom plate 7, and the sample pad 1. Between the gold label pad 2, the nitrocellulose membrane 3 and the water-absorbing pad 6, contact and partially overlap with and only with adjacent parts in turn; the nitrocellulose membrane 3 is adhered to the middle part of the bottom plate 7 , which is provided with a detection line 4 and a quality control line 5 at intervals, the detection line 4 is close to the gold standard pad 2, and the quality control line 5 is close to the water absorption pad 6;

[0122] Rabbit anti-dog IgG labeled with colloidal gold is coated on the gold...

Embodiment 3

[0145] Embodiment 3 A kind of rabies antibody detection kit and detection method

[0146] This example provides a rabies antibody detection kit, including: the purified rabies virus recombinant antigen in Example 1, the concentration is 0.5 μg / mL.

[0147] Further, the enzyme-labeled secondary antibody is also included, and the dilution of HRPO (horseradish peroxidase)-labeled SPA (Staphylococcus protein A) is 1:10000.

[0148] Further, a blocking solution is also included, which is 100 g of Tris, 50 g of casein, and 500 g of sucrose, added with water to 30 L, pH 7.5.

[0149] Further, it also includes sample diluent, PBS buffer containing 10% bovine serum.

[0150] Further, it also includes TMB substrate solution.

[0151] The present embodiment provides an indirect ELISA method for detecting rabies antibodies using the above-mentioned kit, comprising the following steps:

[0152] Coating: The rabies virus recombinant antigen sample (1.0mg / mL~2mg / ml) purified in Example 1 ...

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Abstract

The present invention relates to the technical field of rabies application, in particular to a rabies virus recombinant antigen and a preparation method and use thereof, including one of the following (a)-(c): (a), in the direction from the N-terminal to the C-terminal, including Such as amino acid sequence fragments N, G and P; (b), amino acid sequence fragments N, G and / or P in (a) have been substituted, deleted or added at least one or several amino acid recombinant antigens of rabies virus, and can be Inducing an immune response in a mammal against the rabies virus recombinant antigen; or (c), an amino acid sequence having homology to the amino acid sequence of the rabies virus recombinant antigen in (a), and eliciting a mammal against the rabies virus Immune response of recombinant antigens; the above-mentioned rabies virus recombinant antigens have antigenic epitopes with good antigenicity and non-specificity, and have the advantages of stability, safety, low culture cost and high efficiency compared with holovirus antigens.

Description

technical field [0001] The invention relates to the technical field of rabies application, in particular to a rabies virus recombinant antigen and a preparation method and use thereof. Background technique [0002] Rabies is a fatal and highly neurotropic zoonotic infectious disease caused by rabies virus (RV), which spreads all over the world. Rabies virus can cause fatal central nervous system infection in humans and various animals. The main clinical manifestations are hydrophobia, photophobia, dysphagia, mania, etc. The fatality rate is almost 100%. According to a report released by the World Health Organization in June 2018, rabies kills 59,000 people each year (nearly half of them are children), accounting for 59.6% in Asia and 36.4% in Africa. 99% of human rabies is transmitted by dogs and cats. Since there is no effective treatment for rabies at present, rabies is mainly controlled by vaccine immunization, and it is particularly important to monitor the antibody lev...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K19/00C12N15/62C07K16/10C07K16/06G01N33/569G01N33/558A61K39/205A61P31/14
CPCC07K14/005C07K16/10C07K16/06G01N33/56983G01N33/558A61K39/12A61P31/14C12N2760/20122C12N2760/20134C07K2319/40G01N2333/145G01N2469/20A61K2039/552Y02A50/30
Inventor 刘巧荣陈西钊马君
Owner 北京世纪元亨动物防疫技术有限公司
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