摘要
为建立鹅圆环病毒(GoCV)抗体的检测方法,本研究通过原核表达重组核衣壳(Cap)蛋白,将纯化的该重组蛋白作为ELISA检测的包被抗原,利用交叉反应性,以羊抗鸡IgG(HRP-IgG)作为二抗,经反应条件的优化,建立了检测鹅血清GoCV抗体的间接ELISA方法。确立的ELISA反应最适条件为:抗原包被浓度为2.05μg/mL,待检血清稀释度为1∶50,HRP-IgG稀释度为1∶2000,待测血清和二抗均于37℃反应1.5h,底物于室温显色10 min。经38份阴性血清检测确定阴阳性临界值为0.25。特异性试验结果显示,该方法与大肠杆菌、新城疫、禽流感H5和H9、小鹅瘟、鹅副粘病毒抗原阳性血清均无交叉反应;重复性试验结果显示,其批内和批间变异系数均小于10%。应用该方法检测了从4群GoCV PCR检测阳性种鹅群采集的血清,结果显示其血清阳性率在60.0%～90.9%。本实验建立的间接ELISA方法具有较好的特异性和重复性,为进一步开展GoCV流行病学调查提供了一种便捷可靠的手段。
In the present study, an indirect ELISA method was established for rapid detection of antibodies against goose circovirus （GoCV） by coating with GoCV Cap protein expressed in E.coli. Then the reaction conditions were optimized including the concentration of coating antigen, the dilution of testing serum and the reaction time of HRP conjugated anti-chick IgG. The cut off-value was determined as 0.246. A total of 44 serum samples from four geese herds were tested using this method. The result showed that 60.0% to 90.9 % samples were positive. The results indicated that the established indirect ELISA is sensitive and specific, which providing an useful tool for epidemiological investigation of goose circovirus infection.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2013年第2期134-137,共4页
Chinese Journal of Preventive Veterinary Medicine
基金
国家自然科学基金(307716080)
浙江省重点科技项目(2007C22051
011102120)
