基于VP6蛋白的牛轮状病毒抗体间接ELISA检测方法的建立与应用  

Establishment and Application of Indirect ELISA Method for Detection of BRV Antibody Based on VP6 Protein

作　　者：刘广阔 邹敏 吴发兴[3] 于皓同 王凯茸 张锐铮 张琪[1] 许信刚[1] LIU Guang-kuo;ZOU Min;WU Fa-xing;YU Hao-tong;WANG Kai-rong;ZHANG Rui-zheng;ZHANG Qi;XU Xin-gang(College of Veterinary Medicine,Northwest A&F University,Yangling,Shaanxi 712100,China;State Key Laboratory of Animal Genetical Engineered Vaccine of Ministry of Science and Technology,Qingdao YeBio Biological Engineering Company Limited,Qingdao,Shandong,266114,China;National Animal Serum Bank,China Animal Health and Epidemiology Center,Qingdao,Shandong,266114,China)

机构地区：[1]西北农林科技大学动物医学院,陕西杨凌712100 [2]青岛易邦生物工程有限公司/科技部动物基因工程疫苗国家重点实验室,山东青岛266114 [3]中国动物卫生与流行病学中心/国家动物血清库,山东青岛266114

出　　处：《动物医学进展》2024年第1期1-6,共6页Progress In Veterinary Medicine

基　　金：陕西省重点研发计划重点项目(2022NY-098)。

摘　　要：为了建立牛轮状病毒(BRV)血清抗体的检测方法,从病料中克隆牛轮状病毒VP6基因,构建其表达载体,利用原核表达技术表达重组VP6蛋白,建立牛轮状病毒血清抗体间接ELISA检测方法。结果显示,重组VP6蛋白大小为40 ku,以包涵体形式表达,Western blot证实重组VP6蛋白有良好的反应原性;以4μg/mL浓度的抗原包被酶标板,一抗血清50倍稀释,酶标二抗稀释10000倍稀释,为最佳工作条件,阴阳临界值为0.233,表明该方法具有良好的特异性和敏感性。建立的检测BRV抗体的间接ELISA可用于临床BRV感染的检测。In order to establish a method for the detection of bovine rotavirus(BRV)serum antibody,bovine rotavirus VP6 gene was cloned from the diseased material and its expression vector was constructed.The recombinant VP6 protein was expressed by prokaryotic expression method.An indirect ELISA method for detection of bovine rotavirus serum antibodies was established.The results indicated that the size of the expressed recombinant VP6 protein was 40 ku,and the expression form was inclusion bodies.Western blot indicated that the recombinant protein had good reactivity.The resulet also indicated that the optimal working conditions were coated with 4μg/mL antigen,50-fold dilution of the first antiserum and 10000-fold dilution of the enzyme-labeled second antibody.The critical value of negative and positive samples was 0.233.The method has good specificity and sensitivity.This study established an indirect ELISA method for the detection of BRV antibody,which can be used for clinical detection of BRV infection.

关 键 词：牛轮状病毒 VP6蛋白 原核表达 间接酶联免疫吸附试验 

分 类 号：S853.74[农业科学—临床兽医学] S852.42[农业科学—兽医学]