首批Vero细胞蛋白质对照品的研制

Establishment of the first batch of Vero cell protein reference substance

目的建立首批Vero细胞蛋白质国家对照品,用于人用狂犬病疫苗Vero细胞宿主蛋白残留量检测试验的验证。方法本对照品系采用狂犬病病毒固定株接种Vero细胞,经培养、收获、浓缩、灭活病毒、纯化后,加入适量的人血白蛋白、右旋糖酐40冻干制成。本对照品蛋白含量溯源于Vero细胞蛋白质标准品(250022-201901),应用Vero细胞分泌型蛋白检测试剂盒(ELISA)对其进行标定后赋值。随机选取16支标准品,测定其pH值,采用方差分析统计检验均匀性;随机选取对照品,分别在4、25和37℃放置不同时间(1、2、3和4周),不同时间点取3支,以Vero细胞蛋白质标准品(250022-201901)绘制标准曲线,ELISA法测定蛋白含量,采用方差分析统计检验稳定性。结果经协作标定,本对照品标示量值为7.7μg/mL,参考范围为(7.7±3.6)μg/mL,均匀性及稳定性良好。结论Vero细胞蛋白质对照品已获批使用,批号为250023-201901,该对照品的建立,对加强人用狂犬病疫苗的质量控制具有重要意义。

Objective To establish the first batch of Vero cell protein reference substance used for verification of detection test for residual Vero cell host protein.Methods The fixed strain of rabies virus was inoculated into Vero cells,then cultured,harvested,concentrated,inactivated,purified,added with human albumin and dextran 40,and lyophilized.The protein content of the prepared reference substance was traced with Vero cell protein standard(290022-201901),and assigned by calibration with ELISA kit for secretory Vero cell protein.Sixteen containers of standard were randomly selected and determined for pH value,of which the uniformity was verified by analysis of variance.The reference substance was stored at 4,25 and 37℃for various time durations(1,2,3 and 4 weeks),from which three containers were taken at each time points and determined by ELISA,of which the protein content was calculated according to the standard curve plotted with Vero cell protein standard(290022-201901),and the stability was verified by analysis of variance.Results Collaborative calibration showed that the stated value of Vero cell protein reference substance was 7.7μg/mL,of which the reference range was(7.7±3.6)μg/mL.This batch of reference substance showed good uniformity and stability.Conclusion Vero cell protein reference substance has been approved for use,of which the batch number is 250023-201901.The establishment of this reference substance is of a great significance in improving the quality control of rabies vaccine for human use.