中国兽医科学2024，Vol.54Issue(8)：1050-1055,6.DOI:10.16656/j.issn.1673-4696.2024.0137
非洲猪瘟病毒CD2v阻断ELISA检测方法的初步建立与应用
Preliminary establishment and application of a blocking ELISA based on CD2v protein of African swine fever virus
摘要
Abstract
In order to establish a blocking enzyme-linked immunosorbent assay(ELISA)method for specific detection of CD2v antibodies against African swine fever virus(ASFV),CD2v eukaryotic protein was used as coating antigen,and a monoclonal antibody against the CD2v protein of ASFV was used as blocking antibody.After optimization of the reaction conditions,sensitivity and specificity test,a specifically blocking ELISA method was developed.The results showed that the optimal antigen coating concentration was 20 ng/well,the optimal serum dilution was 1∶1,and the optimal dilution of the enzyme-labelled antibody was 1∶10 000.The prelimary judge criteria of blocking ELISA method was as follows:the cut off values of average percentage inhibition(PI)≥41.23％was taken as CD2v antibody positive result,the PI value＜41.23％was CD2v antibody negative result.The specificity test showed that this method had no cross-reactions with positive sera with pseudorabies virus,porcine circovirus 2,porcine reproductive and respiratory syndrome virus,porcine parvovirus and classical swine fever virus,respectively.The sensitivity test showed that a 128-fold dilution of positive serum can be detected by this method.The intra-assay and inter-assay variation coefficients were both less than 10％.The coincidence rate of this method to 64 pig sera was 96.88％.In conclusion,the blocking ELISA based on ASFV CD2v protein established in this study can be applied to the identification of low-virulent ASFV strain with CD2v deletion combining with the serological diagnosis method of p30 protein and other antigens of ASFV.This method provides an effective detection technology for the epidemiological investigation and epidemic monitoring of low-virulent ASFV strain with CD2v deletion in China.关键词
非洲猪瘟病毒/CD2v蛋白/阻断ELISA/敏感性/特异性Key words
African swine fever virus(ASFV)/CD2v protein/blocking enzyme-linked immunosorbent assay/sensitivity/specificity分类
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王彩霞,林祥梅,于浩洋,吴佳俊,杨振,贾红,仇松寅,刘晓飞,吴绍强,冯春燕..非洲猪瘟病毒CD2v阻断ELISA检测方法的初步建立与应用[J].中国兽医科学,2024,54(8):1050-1055,6.基金项目
"十四五"国家重点研发计划项目(2022YFD1800500) （2022YFD1800500）
