荧光素酶免疫吸附法高灵敏检测蓝舌病病毒抗体方法的建立  

Development of an ultrasensitive detection method for anti-BTV antibody based on the luciferase immunosorbent assay

作　　者：张鸿歌 田占成[1] 独军政[1] 王琼洁 康棣 户鑫兵 张忠辉 关贵全[1] 殷宏[1,2] ZHANG Hongge;TIAN Zhancheng;DU Junzheng;WANG Qiongjie;KANG Di;HU Xinbing;ZHANG Zhonghui;UAN Guiquan;YIN Hong(State Key Laboratory for A nimal Disease Control and Prevention/Lanzhou Veterinary Research Institute,Chinese Academy of A gricultural Sciences,Lanzhou 730046,China;Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses,Yangzhou 225009,China)

机构地区：[1]中国农业科学院兰州兽医研究所动物疫病防控全国重点实验室,甘肃兰州730046 [2]江苏省动物重要疫病与人兽共患病防控协同创新中心,江苏扬州225009

出　　处：《中国兽医科学》2024年第7期881-887,共7页Chinese Veterinary Science

基　　金：国家重点研发计划项目(2021YFD1800500);国家肉牛牦牛产业技术体系专项(CARS-37);甘肃省自然科学基金项目(22JR5RA029);甘肃省基础研究创新群体项目(22JR5RA024);甘肃省科技厅项目(22CX8NA011);农业科技创新工程项目(CAAS-ASTIP-2016-LVRI)。

摘　　要：为了建立一种高效检测动物血清中蓝舌病病毒(BTV)抗体的检测方法,以BTV血清群特异性VP7抗原与荧光素酶蛋白相融合表达的标记蛋白(VP7-NLuc)为检测抗原建立了荧光素酶免疫吸附(VP7-NLuc-LISA)方法,用于检测BTV抗体。结果表明:BTV阳性血清能特异性识别VP7-NLuc融合蛋白;监测BTV-1灭活疫苗免疫绵羊血清中BTV抗体动态结果表明,VP7-NLucLISA方法与商品化竞争性ELISA(c-ELISA)试剂盒的检测结果呈高度相关性(R=0.76,P<0.001);对264份田间血清样本的调查结果表明,建立的VP7-NLuc LISA方法与商品化c-ELISA试剂盒的符合率为98.86%,其相对特异性为99.52%,相对敏感性为96.49%。结论:所建立的VP7-NLuc-LISA方法可作为大规模动物BTV流行病学调查和监测的新方法。To establish an efficient method for detecting anti-bluetongue virus(BTV)antibody in animal serum,BTV VP7 protein fused with a reporter Nano-luciferase(VP7-NLuc LISA)as the diagnostic antigen was used for the development of a luciferase immunosorbent assay(LISA).Results showed that BTV-positive serum could specifically recognize VP7-NLuc fused protein.The dynamic monitoring of BTV antibody in serum of sheep immunized with BTV-1 inactivated vaccine showed that the detection results of VP7-NLuc LISA method and commercial competitive ELISA(c-ELISA)kit were highly correlated(R=0.76,P<0.001).In investigating 264 field serum samples,the relative specificity and sensitivity of the VP7-NLuc LISA were 99.52%and 96.49%,respectively,which was 98.86%in agreement with the commercial c-ELISA kit.In conclusion,the established VP7-NLuc-LISA method can serve as a new method for large-scale animal BTV epidemiological investigation and monitoring.

关 键 词：蓝舌病病毒 VP7蛋白 荧光素酶免疫吸附试验 C-ELISA 血清学检测 

分 类 号：S852.65[农业科学—基础兽医学]