【摘要】 目的建立狂犬病病毒抗原的定量检测方法,用于疫苗生产过程中病毒含量的监测。方法用狂犬病病毒aG株免疫豚鼠,制备抗狂犬病病毒多克隆抗体,纯化后以辣根过氧化物酶进行标记。采用双抗体夹心ELISA法建立检测系统。以狂犬病疫苗国家参考品为定量标准,建立剂量反应曲线,确定该方法的灵敏度,并对该方法的精密性、特异性和适用性进行验证。结果制备的多克隆抗体高峰效价为1∶105,纯化后的抗血清蛋白含量为6.45mg/ml。建立的ELISA方法对狂犬病病毒抗原的最低检出量为5.3mIU/ml,最佳定量区间为10～110mIU/ml,相关系数为0.9983,精密性和特异性较好。用该方法对狂犬病疫苗进行抗原定量,与NIH法测定的疫苗效力具有一定的相关性;检测市售的不同毒株和细胞系生产的狂犬病疫苗,其结果在2.4～9.9IU/ml之间。结论已建立了狂犬病病毒抗原的定量检测方法,可对来自不同毒株和不同细胞系的狂犬病疫苗进行快速定量检测,为疫苗生产中的质量控制提供了简便的监测手段。更多还原

【Abstract】 Objective To develop a quantitative assay for rabies virus antigen,which can be used for monitoring of virus con-tent during vaccine production.Methods Polyclonal antibody was prepared by immunizing guinea pigs with rabies virus strain aG,then purified and labeled with horseradish peroxidase(HRP)to develop a double antibody sandwich ELISA.A dose-response curve was plotted by using national reference for rabies vaccine as a quantitative standard.The sensitivity,precision,specificity and feasi-bility of the developed ELISA were evaluated.Results The prepared polyclonal antibody reached a maximum titer of 1 ∶ 105,and an antiserum protein content of 6.45 mg / ml after purification.The developed ELISA showed good precision and specificity,and its minimum detection limit,optimal quantitative range and correlation coefficient were 5.3 mIU / ml,10 ～ 110 mIU / ml and 0.998 3 respectively.The virus antigen content in rabies vaccine determined by the developed ELISA showed a certain correlation to the vac-cine potency determined by NIH method.The determination results of rabies vaccine prepared with various strains and cell lines by the developed ELISA were 2.4 ～ 9.9 IU / ml.Conclusion A quantitative assay for rabies virus antigen is developed,which may be used for the rapid quantitative determination of rabies vaccine prepared with various strains and cell lines and provides a simple monitoring method for quality control during vaccine production.更多还原