基于p72蛋白的非洲猪瘟病毒抗体检测试纸研制  

Development of Immunochromatographic Strip for Detecting the African Swine Fever Virus Antibodies Based on p72 Protein

作　　者：孙亚宁[1] 卢清侠[1] 邢云瑞 杨苏珍[1] 范璐[1] 乔松林[1] 张改平[1] SUN Yaning;LU Qingxia;XING Yunrui;YANG Suzhen;FAN Lu;QIAO Songlin;ZHANG Gaiping(Key Laboratory of Animal Immunology,Henan Academy of Agricultural Sciences,Zhengzhou450002,China)

机构地区：[1]河南省农业科学院动物免疫学重点实验室,河南郑州450002

出　　处：《河南农业科学》2023年第10期120-130,共11页Journal of Henan Agricultural Sciences

基　　金：“中原英才计划”(育才系列)-中原青年拔尖人才项目(ZYYCYU202012148);河南省重点研发与推广专项(科技攻关)(222102110413);河南省农业科学院优秀青年科技基金项目(2022YQ25);河南省自然科学基金青年基金项目(232300421274)。

摘　　要：为了研制灵敏、准确、特异及稳定的非洲猪瘟抗体快速检测试纸产品,以非洲猪瘟病毒(ASFV)p72蛋白为检测抗原,以金黄色葡萄球菌A蛋白(SPA)为拦截线,对比重组蛋白p72的不同状态对试纸检测性能影响,优化标记条件、喷膜浓度、样品垫缓冲体系及成分、显色时间等因素,确定ASFV抗体检测试纸产品化的具体参数,并对试纸的特异性、均一性、准确性及稳定性进行鉴定。结果显示,试纸的检测敏感性为1∶51200,优于商业化ASFV检测试剂盒,与其他猪源病毒阳性血清无交叉反应,变异系数小于10%,在临床样本检测中未见假阴性及假阳性结果,具有良好的敏感性、特异性、均一性及准确性;经加速稳定试验验证,可以室温保存1 a以上;与进口商品化试剂盒的总符合率为91.6%。综上,成功研制了ASFV抗体检测试纸,且试纸的检测性能良好,可以用于ASFV抗体的快速筛查。In order to establish a sensitive,accurate,specific and stable African swine fever virus(ASFV)antibody rapid detection test strip product,this study used ASFV p72 protein and SPA as detection antigen and test line respectively,compared the effects of different states of p72 on the detection performance of the test strip and optimized the conditions,such as labeling conditions,coating concentration of nitrocellulose membrane,sample pad buffer system and components,and coloring time.The specificity,uniformity,accuracy,and stability of the immunochromatographic strip were identified.The results showed that the detection sensitivity of the strip was 1∶51200,which was better than commercial ASFV‑ELISA kits.No cross‑reaction with positive sera of other swine derived viruses was detected,and the coefficient of variation was less than 10%.No false negative and false positive results in clinical sample detection were observed.Therefore,the strip showed good performance in sensitivity,specificity,uniformity and accuracy.The accelerated stability experiment proved that the strip could be stored at room temperature for more than one year,and the total coincidence rate with the imported commercial ELISA kit was 91.6%.In this study,immunochromatographic strip for detecting the ASFV antibodies was successfully prepared,providing a reliable tool in rapid screening of antibodies against ASFV.

关 键 词：非洲猪瘟 p72蛋白 抗体 试纸 优化 

分 类 号：S854.4[农业科学—临床兽医学]