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Competitive Alpha LISA (linked immuno sorbent assay) detection kit for porcine circovirus (PCV) 2 antibody and detection method thereof

A porcine circovirus and detection kit technology, which is applied in the field of immunological detection of porcine circovirus type 2 antibodies, can solve the problems of large differences in specificity and high requirements, and achieves good specificity, strong anti-quenching ability, The effect of short detection time

Inactive Publication Date: 2014-01-08
INSPECTION & QUARANTINE TECH CENT OF CHONGQING ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] The existing detection methods for the disease mainly include virus isolation and identification, immunoperoxidase technology, in situ hybridization (ISH), immunocolloidal gold technology, immunofluorescence test, immunohistochemical technology, antigen capture ELISA, PCR and real time PCR, etc., each has its advantages and disadvantages, and the specificity varies greatly
At the same time, the method has higher requirements on samples and personnel
However, there is no report on the kit and detection method for detecting porcine circovirus type 2 antibody using AlphaLISA technology at present

Method used

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  • Competitive Alpha LISA (linked immuno sorbent assay) detection kit for porcine circovirus (PCV) 2 antibody and detection method thereof
  • Competitive Alpha LISA (linked immuno sorbent assay) detection kit for porcine circovirus (PCV) 2 antibody and detection method thereof
  • Competitive Alpha LISA (linked immuno sorbent assay) detection kit for porcine circovirus (PCV) 2 antibody and detection method thereof

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Experimental program
Comparison scheme
Effect test

Embodiment

[0027] 1. Construction and identification of PCV Cap recombinant protein prokaryotic expression plasmid

[0028] 1. Target fragment amplification

[0029] 1.1, PCR amplification

[0030] Using the genome of strain PCV2 (primers were designed with the reference sequence GenBank No.: NC_005148.1) as a template, carry out polymerase chain reaction (PCR) amplification of the desired fragment PCV Cap gene, its nucleotide sequence is shown in SEQ ID As shown in No.2, the encoded amino acid sequence is shown in SEQ ID No.1.

[0031] Kpn I and Xho I restriction sites were introduced into the upstream and downstream primers respectively. The primers were designed using Primer Preier5 (a primer design software), and the primers were synthesized by Baobiology (Dalian) Co., Ltd.

[0032]Table 1 Primer sequences

[0033]

[0034] The PCR reaction system is shown in Table 2 (total volume 25 μL)

[0035] Table 2 PCR reaction system

[0036] reaction system Volume (μL) ...

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Abstract

The invention discloses a competitive Alpha LISA (linked immuno sorbent assay) detection kit for a porcine circovirus (PCV) 2 antibody and a detection method thereof. The detection kit comprises donor microspheres, receptor microspheres, a PCV 2 Cap protein monoclonal antibody and a Cap protein antigen with a His label. A competitive Alpha LISA detection method for the PCV antibody is created by optimizing test reaction conditions such as the donor microspheres, the receptor microspheres, the monoclonal antibody, the antigen and serum. The kit for detecting the PCV antibody is good in specificity, high in sensitivity, low in usage amount of the serum, low in detection cost and short in detection time, does not need to be washed and can not be influenced by hemolysis.

Description

technical field [0001] The invention relates to immunological detection of porcine circovirus type 2 antibody, in particular to a porcine circovirus type 2 virus antibody competition AlphaLISA detection kit and detection method. Background technique [0002] Porcine circovirus (Porcine circovirus, PCV) belongs to the Circoviridae genus Circovirus. It is a single-stranded negative-strand DNA virus without an envelope. It is one of the smallest known animal viruses and is mainly divided into 2 genotypes. , namely PCV1 and PCV2. PCV1 is non-pathogenic and widely exists in pig herds. PCV2 infection can cause a variety of disease syndromes, such as multisystem failure syndrome in weaned piglets, reproductive failure in pregnant sows, respiratory disease in weaned pigs and finishing pigs, porcine dermatitis and nephritic syndrome, congenital tremor type A 2 in young piglets, etc. disease. PCV2 is a pathogen that was first isolated from sick pigs with multisystemic wasting syndr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577
CPCG01N33/56983G01N33/54313G01N33/561G01N2333/015G01N2469/20
Inventor 聂福平王昱杨俊李应国向海洋王国民肖进文
Owner INSPECTION & QUARANTINE TECH CENT OF CHONGQING ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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