猪圆环病毒2型IgM抗体间接ELISA的建立

猪圆环病毒2型(PCV2)是断奶仔猪多系统衰竭综合症的重要原发性病原,其感染的早期检测有助于及时采取防控措施。以重组衣壳蛋白(Cap蛋白)作为包被抗原,通过对反应条件的优化,建立了PCV2 IgM抗体间接ELISA,其最适抗原包被浓度为1.25 μg／mL,最适血清稀释度为1∶100,临界值为0.35。该检测方法与其他5种常见猪疫病阳性血清无交叉反应,特异性良好。批内、批间重复试验的变异系数均在2%~6%,重复性好。对100份临床样本的检测结果表明,该检测方法与国外同类试剂盒相比,敏感性为90.3%,特异性为92.8%,总符合率为92%。试验结果表明,所建立的PCV2 IgM间接ELISA特异性好和敏感性高,适用于PCV2感染早期的流行病学调查。

Development of Indirect ELISA for Detection of IgM Antibody to PCV2

Postweaning multisystemic wasting syndrome of swine (PMWS)is associated with porcine circovirus type 2(PCV2). Simple and reliable diagnostic methods are needed to detect antibodies against PCV2 for monitoring early PCV2 infection.An indirect enzyme linked immunosorbent assay (ELISA) method based on purified recombinant capsid protein (Cap) was established to detect IgM antibodies against PCV2. The purified PCV2 Cap protein was used to coat 96 well plate as the antigen,and the following factors affected the experiments were optimized. The results demonstrated that optimal concentration of Cap protein for coating was 1.25 μg/mL,the optimal dilution of serum sample was 1∶100. The cutoff value was 0.35. The results showed no cross-reactivity with positive serum of other five porcine virus. The CV of intra-assay and interassay ranged from 2% to 6%. The indirect ELISA was used to detect 100 field serum samples, compared with control IgG/IgM kit,the diagnostic sensitivity,specificity,and accuracy of PCV2 IgM were 90.3%,92.8% and 92%. The result showed that the indirect ELISA was not only sensitive and specific,but also suitable for large scale epidemiological investigation of early PCV2 infection.