Preparation and Identification of Polyclonal Antibody Against the Cap Protein of Porcine Circovirus Type 2

doi:10.13560/j.cnki.biotech.bull.1985.2019-0205

Abstract

Abstract: This experiment was aimed to prepare polyclonal antibodies of Cap protein of porcine circovirus type 2（PCV2）. The gene was amplified from the DNA of PCV2 CAU0673 strain by PCR,whose product was approximately 702 bp. The recombined plasmid pET30a-PCV2-Cap was expressed in E.coli BL21（DE3）and was induced by IPTG. The recombinant Cap protein was purified by Ni-NTA,and used as an immunogen for immunization of New Zealand white rabbits by subcutaneous injection in several sites on back to prepare polyclonal antibodies. The specificity of the serum antibody was determined by western blot,IFA,and the serum antibody titer was detected by indirect ELISA. PCR,digestion and sequencing were used to identify positive plasmid. The results showed that the recombined plasmid pET30a-PCV2-Cap was successfully constructed. The results of SDA-PAGE showed that the recombined protein was successfully expressed in E.coli BL21;（DE3）with a relative molecular weight of 34 kD,mainly existed in a form of inclusion body. The results of western blot showed that this recombined Cap protein was specifically reacted with PCV2 positive pig serum. The prepared rabbit antisera was specifically reacted with the recombined Cap protein and whole virus antigen,and the serum antibody titer was 1∶12 800.

Key words: Porcine circovirus type 2, Cap protein, polyclonal antibody, identification

OU Yun-wen, DAI Jun-fei, MA Bing, ZHANG Jie, DENG Hui-xiang, LI Xiao, ZHANG Xin-ming. Preparation and Identification of Polyclonal Antibody Against the Cap Protein of Porcine Circovirus Type 2[J]. Biotechnology Bulletin, 2019, 35(8): 226-231.

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