猪细小病毒病血清抗体VP2-ELISA检测方法的建立

Establishment of the Indirect ELISA for the Detection of Viral structural protein Antibodies against Porcine Parvovirus

利用纯化的猪细小病毒VP2蛋白为抗原,建立检测猪细小病毒血清抗体的间接VP2-ELISA。最佳反应条件为:抗原包被浓度300 ng/孔,4℃过夜,待检测血清1:50稀释。与猪细小病毒病阳性血清呈阳性反应,与猪瘟、猪伪狂犬病、猪繁殖与呼吸综合征、猪日本脑炎和猪圆环病毒病等5种疾病的阳性血清均无交叉反应。批间、批内试验变异系数均小于10%。用该方法与Ceditest PPV猪细小病毒抗体检测试剂盒对102份血清进行平行检测和比较,间接VP2-ELISA的敏感性为93.8%,特异性为81.1%,符合率为89.2%。结果表明,猪细小病毒血清抗体间接VP2-ELISA检测方法具有较高的敏感性和特异性,且重复性好,可用于猪细小病毒感染的血清抗体检测。

In order to detect porcine parvovirus antibodies, an indirect ELISA was established, by using the recombinant VP2 protein as antigen. The reaction conditions of the VP2-ELISA were explored and optimized. The optimal coating concentration was 300 ng per well, the coating condition was at 4℃ overnight, and serum samples were diluted to 1：50. The results showed the VP2-ELISA assay was able to detect PPV positive serum, and had no cross-reactivity with serum against HCV, PRV, PRRSV, JEV and PCV, respectively. The intra and inter-assay coefficients of variation （CV） of the VP2-ELISA were all less than 10%. 102 swine serum samples were detected by the VP2-ELISA and Ceditest PPV ELISA Kit in parallel. The sensitivity and specificity of the VP2-ELISA were 93.8% and 81.1% respectively. The coincidence rate between these two assays was 89.2%. The results indicated that the indirect VP2-ELISA was highly sensitive and specific, and could be used for clinical detection of PPV antibodies.