猪流行性乙型脑炎病毒prM-E抗原捕获ELISA检测方法的建立  

Development of an antigen capture ELISA for detecting the prM-E protein of porcine Japanese encephalitis virus

作　　者：王晓磊[1] 霍红[1] 郭立平[1] 李伟[1] 步志高[1] 华荣虹[1] 

机构地区：[1]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/农业部兽医公共卫生重点开放实验室,黑龙江哈尔滨150001

出　　处：《中国预防兽医学报》2015年第6期465-468,485,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家公益性行业(农业)科研专项(201203082);哈尔滨市创新人才项目(2013RFQYJ006);中央级公益性科研院所基本科研业务费专项

摘　　要：为建立猪流行性乙型脑炎病毒(JEV)的检测方法,本研究以pr M-E结构蛋白特异性单克隆抗体(MAb)5B10和5E7分别作为捕获抗体和辣根过氧化酶标记的检测抗体,对反应条件进行优化,建立了抗原捕获ELISA(AC-ELISA)方法。该方法与其它常见猪病病原无交叉反应,特异性强。该方法可以检出1.25×105 pfu/m L的病毒和48.44 ng/m L pr M-E重组蛋白,最佳线性检测范围为0.05μg/m L^1.00μg/m L,线性相关系数(R2)为0.996。组装的试剂盒批内和批间重复性试验的变异系数均小于10%,具有良好的重复性。试剂盒4℃保存12个月稳定。该研究为JEV亚单位疫苗的研制与生产提供了技术支持。To develop a method for detecting porcine Japanese encephalitis virus （JEV）, an antigen capture ELISA （AC-ELISA） was established with monoclonal antibody （MAb） 5B10 as capture antibody and horseradish peroxidase conjugated MAb 5E7 as detection antibody. The AC-ELISA was specificity which had no cross-reactivity with other related pathogens of swine diseases. The detection limit was 1.25 ×105 pfu/mL for JEV and 48.44 ng/mL for viral prM-E protein expressed by mammalian cell lines. The linear quantitative range was 0.05 μg/mL to 1.00 μg/mL, and the liner correlation （R2） was 0.996. The coefficients of variation intra-assay and inter-assay were less than 10%. The AC-ELISA kit was validity for 12 months stored at 4 ℃. The established AC-ELISA kit provided technical support to the development and production of JEV subunit vaccine.

关 键 词：流行性乙型脑炎病毒 prM-E 抗原捕获ELISA 检测 

分 类 号：S852.65[农业科学—基础兽医学]