肠道病毒71型灭活疫苗滴度的酶联免疫吸附检测方法

An ELISA method for the titer determination of enterovirus 71 inactivated vaccine

目的 建立肠道病毒71型（EV71）灭活疫苗滴度的检测方法,用于疫苗研制及生产过程中抗原含量的测定.方法 以兔抗EV71多克隆抗体为包被抗体,小鼠抗EV71单克隆抗体为检测抗体,以夹心ELISA方法测量我们制备的EV71疫苗抗原参照品,建立抗原标准曲线,确定定量限度及最佳定量范围,同时验证该方法的特异性和可重复性.结果 建立了检测EV71抗原含量的夹心ELISA方法,定量限度为3.9352log10TCID50/mL,最佳定量范围为3.9352～5.9686 log10TCID50/mL,相关系数为R2=0.9956;特异性好,除与EV71疫苗样品有反应外,与其他样品无交叉反应;稳定性和重复性好,变异系数小于7%.结论 本研究建立的检测EV71灭活疫苗滴度的夹心ELISA方法灵敏、特异,为疫苗研制过程中抗原含量测定提供了快捷有效的检测手段.

Objective To establish a quantitative assay method for determining the titer of enterovirus 71（EV71）during its inactivated vaccine development and manufacturing.Methods By using a mouse anti-EV71 monoclonal antibody and rabbit anti-EV71 polyclonal antibodies,a sandwich ELISA method for EV71 inactivated vaccine was established.The sensitivity,specificity and reproducibility of the method were evaluated.Results A specific and sensitive ELISA method for quantitative assay of EV71 inactivated vaccine was developed.Its detection limit was determined to be 3.9352 log10TCID50/mL and its detection range was 3.9352-5.9686 log10TCID50/mL.Its correlation coefficient was R2 =0.9956.The method showed good sensitivity,specificity and reproducibility.It had no cross-reaction to samples other than EV71 and the variation co-efficient was lower than 7%.Conclusions A quantitative ELISA method is established for EV71 inactivated vaccine.It offers a useful and rapid way to determine the titer of EV71 inactivated vaccine during its developnent and production.