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Development and Application of Indirect ELISA Antibody for Porcine Circovirue Type 2 and Production of Monoclonal Antibodies Against PCV2

Author: QiYanHua
Tutor: WangAiPing；ZhangGaiPing
School: Zhengzhou University
Course: Cell Biology
Keywords: Porcine circovirus type 2 ORF2 gene fragment The prokaryotic expression Indirect ALISA Monoclonal antibodies
CLC: S858.28
Type: Master's thesis
Year: 2010
Downloads: 112
Quote: 1
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Abstract


Weaning multisystemic wasting syndrome (Postweaning multisystemic wasting syndrome, PMWS) is an important infectious disease of pigs caused by porcine circovirus type 2 virus (Porcine circovirus type2, PCV2). PMWS earliest in western Canada in 1991, was found, after it popular worldwide. The disease mainly against weaned piglets, clinical manifestations of the piglets fever, progressive weight loss, pallor, jaundice, respiratory failure, and digestive system disorders and other symptoms. Porcine circovirus (Porcine circovirus, PCV) belongs to the the ring virus family circovirus genus, is a DNA virus. Divided into type 1 (PCV1) and type 2 (PCV2). PCV1 cell contaminants, non-pathogenic; PCV2 pathogenic PCV2 infection can cause immune suppression caused by the secondary and concurrent infection of pig-related diseases, such as proliferative and necrotizing pneumonia (PNP), porcine dermatitis and nephritis syndrome (PDNS), and porcine respiratory syndrome (PRDC), congenital tremor, enteritis, reproductive disorders. Currently, PCV2 has become one of the most serious harm to the pig industry worldwide pathogen. PCV2 genome contains 11 open reading frames, ORF1 and ORF2 reading frame. ORF1 encodes Rep proteins associated with viral replication, and this protein has 85% homology between the two virus, is the main reason for causing the cross-reaction of the PCV1 and PCV2 antigen. ORF2 encodes the Cap protein is the major structural protein of the virus, the type-specific antigens, having a good immunogenicity and antigenic cross-reactivity between the two type PCV does not occur. This study established the PCV2 antibody indirect ELISA. The feature of this method is sensitive, rapid, specific, and be able to effectively identify the type 2 PCV infection has major implications on the the PCV2 rapid diagnosis, epidemic surveillance, prevention and control. Cloned PCV2 ORF2 gene fragment, ORF2 gene fragment expressed in prokaryotic expression system. The results showed that the gene fragment was successfully expressed in E. coli BL21 fusion protein of relative molecular mass of approximately 26kDa, and can be identified for PCV2 positive serum. 2 to Cap protein expression as a package antigen to establish an indirect ELISA for the detection of PCV2 Cap protein; using the established indirect ELISA method detected 80 sera of pig farms, and control with a commercial kit detection The positive rate of 60%, the two methods in accordance with the rate of 93%. 3. PCV2 Henan strains of virus antigen to immunize Balb / c mice, through the establishment of an indirect ELISA for detection screening, the final two secreting anti-PCV2 monoclonal antibody cell lines: 2B11-C1 3F2-A10. Specificity experiments show that: 2B11-C1 3F2-A10 with classical swine fever virus, swine pseudorabies virus, swine Japanese encephalitis virus, porcine reproductive and respiratory syndrome virus and normal PK-15 cell response, specificity was good. PCV2 monoclonal antibody obtained two further establish specific, sensitive, accurate and rapid detection of PCV2 method laid a foundation.

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CLC: > Agricultural Sciences > Livestock, animal medicine,hunting,silkworm,bee > Animal Medicine ( Veterinary Medicine) > Livestock, poultry, wildlife diseases > Livestock > Pig
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