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African swine fever virus mgfs and cd2v ELISA antibody detection kit

A technology of African swine fever virus and kit, which is applied in the direction of virus/bacteriophage, virus, virus peptide, etc., to achieve the effects of strong specificity, less antigen consumption, and improved efficiency

Active Publication Date: 2022-05-06
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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  • African swine fever virus mgfs and cd2v ELISA antibody detection kit
  • African swine fever virus mgfs and cd2v ELISA antibody detection kit
  • African swine fever virus mgfs and cd2v ELISA antibody detection kit

Examples

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Effect test

Embodiment 1

[0043] Embodiment 1, the preparation of coating antigen of African swine fever virus MGF360-505R and CD2v synthetic peptide ELISA antibody detection kit

[0044] 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

[0074] Example 2, Preparation of positive control serum of African swine fever virus MGF360-505R and CD2v synthetic peptide ELISA antibody detection kit

[0075] 1. Immune animals Use healthy pigs with a body weight of about 40kg (the commercial African swine fever virus antigen and antibody detection kits are all negative), and isolate and observe for 7 days.

[0076] 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.

[0077] 3. Immunization procedure Two times of immunization were carried out at week 0 and week 3 respectively. The immunization doses were 2ml and 4ml re...

Embodiment 3

[0079] Example 3, Preparation of African swine fever virus MGF360-505R and CD2v synthetic peptide ELISA antibody detection kit

[0080] Include the following steps:

[0081](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.

[0082] (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).

[0083] (3) Negative control serum: specific pathogen-free (SPF) pig serum, used as the negative control serum of the kit (1 tube, 1.5ml / tube).

[0084] (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 solutio...

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Abstract

The invention discloses an African swine fever virus MGFs and CD2v ELISA antibody detection kit. The kit includes an enzyme-linked reaction plate and an enzyme-labeled secondary antibody, wherein the enzyme-labeled plate is coated with an African swine fever virus antigen epitope polypeptide composition. The antigen epitope polypeptide composition is one or any combination of the polypeptide shown in sequence 1 in the sequence listing, the polypeptide shown in sequence 2 in the sequence listing, and the polypeptide shown in sequence 3 in the sequence listing . This kit uses a chemically synthesized antigen peptide to coat the microtiter plate, with less antigen consumption, high sensitivity and specificity, and can efficiently detect whether there are MGF360-505R and CD2v antibodies produced by wild virus infected with African swine fever virus, so as to realize Differential diagnosis. The kit of the invention has high sensitivity, good specificity, convenient operation and good market prospect.

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] 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 of susceptible pigs is as high as 100%. [0003] ASFV belongs to the African swine fever virus genus (Asfivirus) of the African swine fever virus family (Asfarviridae). ASFV is an enveloped virus. The viral genome is a double-stranded DNA molecule, 170-193kb in length, containing 151-167 open reading frames (ORFs), encoding more than 150 polypeptides, and about 30% of the ORFs have multiple copies, ...

Claims

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Application Information

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Patent Type & Authority Patents(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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