狂犬病病毒糖蛋白抗原含量双抗体夹心ELISA检测方法的建立及验证Development and validation of double antibody sandwich ELISA for determination of rabies virus glycoprotein antigen content
郭笑语;潘东;陈晓旭;韩丹丹;高一峰;沈艳杰;韩尚成;赵天罡;丁楠;张洁琼;袁若森;姜春来;
摘要(Abstract)：
目的建立可快速检测狂犬病病毒(rabies virus,RabV)糖蛋白抗原含量的双抗体夹心ELISA方法,并进行验证及初步应用。方法用抗RabV糖蛋白基因工程抗体建立双抗体夹心ELISA法,检测RabV糖蛋白抗原含量。对建立的方法进行特异性、线性、准确性、板内与板间精密性、灵敏度验证,对不同厂家(毒株分别为aG、PV、CTN、PM)生产的狂犬病疫苗与不同工艺阶段的狂犬病疫苗样品进行适用性检测。结果捕获抗体与酶标抗体最佳稀释度分别为1∶2 000和1∶4 000,最佳封闭剂为1. 5%BSA。该方法与其他人用疫苗和辅料成分无交叉反应;线性范围在0. 003 2～0. 103 1 IU/mL之间,R2> 0. 98;准确性验证的回收率在97. 0%～110. 1%之间;精密性验证的板内变异系数<8. 6%,板间变异系数<13. 9%;检测不同厂家的狂犬病疫苗和不同工艺阶段的疫苗样品呈良好的剂量依赖效应。结论建立了适用于人用狂犬病疫苗糖蛋白抗原含量检测的双抗体夹心ELISA方法,符合定量检测的需求,可用于aG、PV、CTN、PM等不同毒株生产的狂犬病疫苗检测;也可用于病毒收获液、浓缩液、灭活液及原液等不同工艺阶段样品检测。
关键词(KeyWords)： 狂犬病病毒;糖蛋白;抗原;定量检测;酶联免疫吸附测定
基金项目(Foundation):
作者(Authors): 郭笑语;潘东;陈晓旭;韩丹丹;高一峰;沈艳杰;韩尚成;赵天罡;丁楠;张洁琼;袁若森;姜春来;
DOI: 10.13200/j.cnki.cjb.003038
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