ELISA antibody detection kit for African swine fever viruses MGFs and CD2v

A technology of African swine fever virus and kit, applied in the direction of virus/bacteriophage, virus, virus peptide, etc., to achieve the effect of improving efficiency, high sensitivity, and broad market prospects

Active Publication Date: 2020-11-20
CHINA ANIMAL HUSBANDRY IND
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although there is currently no African swine fever vaccine approved for marketing at home and abroad; however, current research shows that the African swine fever gene deletion vaccine is the most likely and most promising African swine fever vaccine, especially MGF360-505R Gene deletion strains or natural attenuated strains of multi-genome families and CD2v have good safety and immune protection effects, and can provide challenge protection against homologous genotypes and heterologous genotypes after immunization

Method used

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  • ELISA antibody detection kit for African swine fever viruses MGFs and CD2v
  • ELISA antibody detection kit for African swine fever viruses MGFs and CD2v
  • ELISA antibody detection kit for African swine fever viruses MGFs and CD2v

Examples

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Embodiment 1

[0044] Embodiment 1, the preparation of coating antigen of African swine fever virus MGF360-505R and CD2v synthetic peptide ELISA antibody detection kit

[0045] In this experiment, bioinformatics methods were used to accurately analyze the main epitopes of MGF360, MGF505 and CD2v proteins in the Pig / HLJ / 2018 strain sequence (MK333180.1) published by GenBank, and to screen out suitable peptides. Three peptides were synthesized by the peptide synthesizer, and the sequences were respectively shown in sequence 1, sequence 2, and sequence 3 in the sequence table, and the coated antigen with a purity of about 80% was produced, which can cover the protein of African swine fever virus MGF360, MGF505 and CD2v The main antigenic epitope increases the detection rate of antibody positive. The polypeptide synthesis method can be a conventional method. The following method is used in the present invention to synthesize the three polypeptides of the present invention as the coating agent of...

Embodiment 2

[0076] Example 2, Preparation of positive control serum of African swine fever virus MGF360-505R and CD2v synthetic peptide ELISA antibody detection kit

[0077] 1. Healthy pigs with a body weight of about 40kg were used as immunized animals (the commercial African swine fever virus antigen and antibody detection kits were all negative), and they were isolated and observed for 7 days.

[0078] 2. Preparation of immunogen Dilute the epitope polypeptide containing sequence 1, sequence 2 and sequence 3 with sterile deionized water (the mass ratio of each antigenic peptide is 1:1:1) to a 2 mg / ml solution, and dilute it with an equal volume Freund's complete adjuvant mixed with Freund's complete adjuvant, fully emulsified for the first immunization, mixed with an equal volume of Freund's incomplete adjuvant, fully emulsified for the second immunization.

[0079] 3. The immunization program is divided into two immunizations, immunizations are carried out at 0 weeks and 3 weeks respe...

Embodiment 3

[0081] Example 3, Preparation of African swine fever virus MGF360-505R and CD2v synthetic peptide ELISA antibody detection kit

[0082] Include the following steps:

[0083] (1) A 96-well detachable polystyrene enzyme-linked reaction plate coated with African swine fever virus MGF360-505R and CD2v epitope polypeptide antigen; 2×96 wells.

[0084] (2) Positive control serum: after the hyperimmune serum prepared by immunizing the epitope polypeptide shown in the African swine fever virus sequence 1, sequence 2 and sequence 3 is diluted with the sample diluent, it is used as the positive control serum of the kit (1 tube, 1.5ml / tube).

[0085] (3) Negative control serum: specific pathogen-free (SPF) pig serum, used as the negative control serum of the kit (1 tube, 1.5ml / tube).

[0086] (4) Enzyme-labeled secondary antibody: prepared by diluting 1:30000 with horseradish peroxidase-labeled rabbit anti-pig IgG (purchased from sigma company, product number A5670) as the stock soluti...

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Abstract

The invention discloses an ELISA antibody detection kit for African swine fever viruses MGFs and CD2v. The kit comprises an ELISA plate and an enzyme-labelled secondary antibody, wherein the ELISA plate is coated with an African swine fever virus epitope polypeptide composition. The epitope polypeptide composition is one or arbitrary combination of two or more of polypeptide represented in the sequence 1 in the sequence table, polypeptide represented in the sequence 2 in the sequence table and polypeptide represented in the sequence 3 in the sequence table. The kit adopts chemically synthesized antigen peptide to coat the ELISA plate, is low in antigen dosage and high in sensitivity and specificity, and can efficiently detect whether MGF360-505R and CD2v antibodies produced by wild strainsinfected with the African swine fever viruses exist or not, and thereby realizing differential diagnosis. The kit is high in sensitivity, good in specificity and convenient to operate and has good market prospects.

Description

technical field [0001] The invention belongs to the technical field of biological detection. More specifically, the invention relates to an African swine fever virus ELISA antibody detection kit, in particular to an African swine fever virus MGF360-505R and CD2v synthetic peptide ELISA antibody detection kit. Background technique [0002] In August 2018, African swine fever broke out in my country. This is the first ASF incidence record in the history of our country. Since then, the prevention and control of African swine fever has been promoted to the primary position of domestic swine diseases, which has brought a devastating blow to the breeding industry. , which has greatly affected the supply of live pigs in our country. [0003] African swine fever is an acute, febrile, highly contagious zoonotic disease of pigs caused by African swine fever virus (ASFV), clinically characterized by high fever, reticuloendothelial hemorrhage, and high mortality , the case fatality rate...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/01G01N33/569G01N33/543G01N33/58G01N33/68
CPCC07K14/005G01N33/56983G01N33/54306G01N33/581G01N33/6854C12N2710/12022G01N2333/01G01N2469/20
Inventor 张蕾董春娜李静毕建敏肖进齐鹏刘新月李玲李鹏宇吴冬荀
Owner CHINA ANIMAL HUSBANDRY IND
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