Hog cholera virus inhibition ELISA antibody detection kit and application thereof

A swine fever virus and antibody detection technology, applied in the direction of virus/bacteriophage, application, virus, etc., can solve the problems of high non-specificity, low negative coincidence rate, long detection time, etc., and achieve high cell culture density, high sensitivity and specific effect

Active Publication Date: 2015-04-01
WUHAN KEQIAN BIOLOGY CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, most of the ELISA methods for detecting swine fever antibodies established in China are indirect ELISA methods. Although the indirect ELISA method for detecting swine fever antibodies appeared earlier and has good sensitivity, it is often inevitable that there will be high non-specificity.
Secondly, most of the coating antigens used in the established ELISA method for detecting classical swine fever antibodies are expressed in prokaryotic systems, but the structural protein E2 of classical swine fever virus is a glycosylated protein, and the prokaryotic expression products cannot be glycosylated and other proteins. Modification, it is quite different from the natural E2 prot

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  • Hog cholera virus inhibition ELISA antibody detection kit and application thereof
  • Hog cholera virus inhibition ELISA antibody detection kit and application thereof

Examples

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Embodiment 1

[0033] A large-scale production of classical swine fever virus blocking ELISA antibody detection kit, the kit includes: using the Bac to Bac baculovirus expression system and the Sf9 cell suspension culture process to express and purify the classical swine fever virus E2 protein as a coated plate Antigen, horseradish peroxidase-labeled monoclonal antibody against CSFV E2 protein, the monoclonal antibody is prepared using expressed and purified CSFV E2 protein as immunogen.

[0034] The kit also includes: antigen-coated plate, positive control, negative control, sample diluent, 20 times concentrated washing solution, chromogenic solution A, chromogenic solution B, stop solution;

[0035] The antigen-coated plate uses 0.1mol / L, pH9.6 sodium bicarbonate solution as the coating liquid, and after diluting the expressed and purified E2 protein to a certain concentration, coats the ELISA plate;

[0036] The positive control is hyperimmune serum immunized with live swine fever vaccine...

Embodiment 2

[0057] A kind of application of swine fever virus blocking ELISA antibody detection kit in detection swine fever virus antibody, its steps are:

[0058] 1) Take out the antigen-coated plate coated with CSFV E2 protein from the kit, and dilute the serum to be tested with the sample

[0059] solution was diluted 1:4, 100 μL per well was added to the antigen-coated plate, and negative and positive control wells were set at the same time, each with 2 wells, 100 μL per well;

[0060] 2) Gently shake the samples in the wells, place at 37°C for 45 minutes, shake off the solution in the wells, wash the plate 3 times with washing solution,

[0061] 200μL per well, pat dry on absorbent paper for the last time;

[0062] 3) Add 100 μL of enzyme-labeled monoclonal antibody to each well, place at 37°C for 30 minutes, and wash 3 times, the method is the same as step (2). 25°C) to develop color in the dark for 10 minutes, add 50 μL of stop solution to each well, and use a microplate reader ...

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Abstract

The invention discloses a hog cholera virus inhibition ELISA antibody detection kit and application thereof. The kit comprises hog cholera virus E2 protein which is expressed by virtue of a baculovirus expression system and a cell suspension culture process; the hog cholera virus E2 protein is purified to serve as a coated plate antigen and an immunogen to prepare a monoclonal antibody of the hog cholera virus E2 protein, and horse radish peroxidase (a hybridoma cell strain 4A7 CCTCC NO.C2014229) is marked. The kit further comprises an antigen coated plate, a positive control, a negative control, sample diluent, a scrubbing solution, a developing solution A, a developing solution B and a stop solution. According to the application of the hog cholera virus inhibition ELISA antibody detection kit in the detection of the hog cholera virus antibody, the kit is used for detecting other positive serums except hog cholera viruses, the cross reaction is avoided, the specificity is strong, the sensitivity is high, the detection time is short, and the repeatability is good; compared with an import kit, the detection coincidence rate reaches above 95%.

Description

technical field [0001] The present invention relates to the detection of animal epidemics, in particular to an enzyme-linked immunoimmunoassay kit capable of large-scale production for detecting antibodies against swine fever virus, and at the same time relates to a detection kit for detecting antibodies against swine fever virus by ELISA in the application. It is suitable for detecting the antibody level of swine fever virus in pig serum. Background technique [0002] Classical Swine Fever (CSF) is one of the infectious diseases that seriously harm the swine industry. It is highly contagious and highly pathogenic, and has caused huge economic losses to the swine industry worldwide. It has been recognized by the World Organization for Animal Health (OIE) is listed as one of the 16 infectious diseases in category A, and it is an animal infectious disease that must be reported. my country also lists it as a first-class infectious disease. In recent years, the symptoms of swin...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/569C07K14/18C07K16/10C12N15/866
CPCC07K14/005C07K16/1081C12N2770/24051G01N33/56983
Inventor 华俊清王艳伟但汉并周明光董晓辉徐高原金梅林陈焕春
Owner WUHAN KEQIAN BIOLOGY CO LTD
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