期刊论文

中文

中国预防兽医学报

1008-0589

2012

34

11

894-897

10.3969/j.issn.1008-0589.2012.11.11

湖南省科技重大专项（2007FJ1003） 湖南农业大学***基金（04YJ03）

本校为第一机构

为建立一种快速检测猪圆环病毒2型(PCV2)抗体的方法,本研究通过基因合成PCV2的Rep编码序列,将其克隆至pET-28a(+)并在E.coli中表达,表达的重组Rep蛋白经Ni~(2+)亲和纯化。采用纯化的目的蛋白作为包被抗原,建立检测PCV2 Rep蛋白抗体的间接ELISA方法。结果表明最佳抗原包被浓度为200 ng、孔,血清和二抗最佳反应时间分别为lh和30min。该方法具有良好的特异性和敏感性,与韩国金诺公司试剂盒的符合率为81.4%,表明建立的ELISA方法可以用于PCV2的流行病学检测。

To develop a method for porcine circovirus 2 （PCV2） detection, the gene coding the Rep of PCV2 was synthesized and cloned into pET-28a（＋）, which was expressed in Escherichia coli. The recombinant Rep protein was purified by nickel affinity chromatography and used as the coating antigen to establish an indirect ELISA as a fast method to detect PCV2 antibody. The reaction conditions were optimized which included 200 ng of recombinant Rep as coating antigen for per well, and an hour and a half incubation for sample serum and secondary antibod...