DEV gB protein monoclonal antibody and blocking ELISA kit for detecting DEV antibody

A detection kit, monoclonal antibody technology, applied in the direction of antibodies, antiviral agents, immunoglobulins, etc., can solve the lack of blocking ELISA kits and other problems, achieve good commercial application prospects, high sensitivity, repeatability strong effect

Active Publication Date: 2020-02-14
HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

So far, there is still a lack of blocking ELISA kits for the detection of DEV antibodies with commercial application value

Method used

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  • DEV gB protein monoclonal antibody and blocking ELISA kit for detecting DEV antibody
  • DEV gB protein monoclonal antibody and blocking ELISA kit for detecting DEV antibody
  • DEV gB protein monoclonal antibody and blocking ELISA kit for detecting DEV antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Example 1 Prokaryotic expression of DEV gB protein

[0054] 1. Materials and methods

[0055] 1.1 Virus strains and experimental animals

[0056] DEVCSC strain seed virus was preserved by our laboratory; SPF duck embryos were purchased from the Experimental Animal Center of Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences; clean grade New Zealand white rabbits were purchased from Liaoning Changsheng Biotechnology Co., Ltd.; DEV positive serum reference materials were purchased from China Veterinary Drug Supervision Institute; pET-32a vector plasmid is preserved by our laboratory.

[0057] 1.2 Test method

[0058] 1.2.1 Proliferation of DEV

[0059] Using conventional methods, duck embryo fibroblasts (DEFs) were prepared from 11-day-old SPF duck embryos, and the prepared primary cells were cultured in DMEM containing 5% FBS, and the primary cells were placed in 5% CO 2 In a 37°C incubator, subculture after 1-2 days, and grow to 80% of th...

Embodiment 2

[0129] Example 2 Preparation and selection of DEV and DEV gB protein monoclonal antibody

[0130] 1. Test material

[0131] 1.1 Virus strains and cells

[0132] H5, H7 and H9 subtype avian influenza virus (AIV) HI test antigens were purchased from Harbin Veken Biotechnology Development Company, Newcastle disease virus (NDV), egg drop syndrome virus (EDSV), duck Tembusu virus (DTMUV) Duck hepatitis virus type Ⅰ (DHVI Ⅰ) are all preserved by our laboratory. SP2 / 0 myeloma cells are maintained by the laboratory.

[0133] 1.2 Experimental animals and serum samples

[0134] Eleven-day-old SPF duck embryos were purchased from the Experimental Animal Center of Harbin Veterinary Research Institute. Six-week-old SPF female BALB / c mice were purchased from Beijing Weitong Lihua Experimental Animal Technology Co., Ltd. 6 positive serum samples were prepared from immunized SPF ducks in the laboratory and identified by neutralization test; 6 negative serum samples were collected from no...

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Abstract

The invention discloses a DEV gB protein monoclonal antibody and a blocking ELISA kit for detecting a DEV antibody. The preparation method comprises the following steps: firstly, preparing gB proteinwith good reactogenicity and immunogenicity through a prokaryotic expression system, wherein the hybridoma cell strain with a microbial preservation number of CGMCC No.17996 is obtained by screening through a cell fusion technology by taking the hybridoma cell strain as an antigen, and the secreted monoclonal antibody can react with duck enteritis virus native space conformation virus protein andhas strong blocking capacity on a DEV antibody. The invention also provides an epitope recognized by the monoclonal antibody, and the amino acid sequence of the epitope is shown as SEQ ID No. 1. The invention further discloses the blocking ELISA kit for detecting the DEV antibody by applying the monoclonal antibody. The blocking ELISA kit has the advantages of high sensitivity, good specificity, strong repeatability and the like.

Description

technical field [0001] The invention relates to a monoclonal antibody and a blocking ELISA kit, in particular to a DEV gB protein monoclonal antibody and a blocking ELISA kit for detecting DEV antibodies, belonging to the field of antibody detection of duck viral enteritis virus. Background technique [0002] Duck virus enteritis (DVE), commonly known as duck plague (Duck plague, DP), is an acute, febrile, septic infectious disease of ducks, geese, wild geese and other birds of Anseriformes. Viral enteritis virus (Duckenteritis virus, DEV) caused. The viral genome encodes more than a dozen glycoproteins, among which the gB protein is one of the main proteins displayed on the surface of the viral envelope and is also the main immunogenic protein. Duck viral enteritis is the herpes virus disease of anatidae, seriously endangers domestic ducks, and also attacks multiple water fowls such as wild ducks. It was diagnosed for the first time in the United States in 1967. At first,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/20C07K16/08C07K14/03G01N33/569A61K39/42A61P31/22C12R1/91
CPCC07K16/085C07K14/005G01N33/56983A61P31/22C12N2710/16022C07K2317/35Y02A50/30
Inventor 赵丽丽陈洪岩王玉娥
Owner HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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