摘要
目的制备病毒载体疫苗滴度测定用国家标准品。方法以鸡胚成纤维细胞制备痘苗病毒,经密度梯度超速离心纯化后,加入天花疫苗保护剂,分装冻存,分发至我国4个不同实验室,按照统一的结晶紫染色蚀斑法实验方案进行协作标定,每次实验稀释后取2个稀释度,每个稀释度设5个平行孔,计算各实验室检测结果的几何均数,对标准品中痘苗病毒滴度进行赋值。检测病毒标准品于不同温度下保存不同时间的稳定性,并对病毒标准品的各项质量指标进行检定。结果 4个实验室共测定44次,4个实验室间的检测结果差异无统计学意义(P>0.05),各实验室检测结果的几何平均数为3.05×106PFU/ml,95%CI为2.00×106～4.65×106PFU/ml,标准差为0.09,总变异系数1.41%。制备的痘苗病毒标准品于4℃可稳定放置10 d,37℃放置稳定性较差。制备的痘苗病毒标准品装量检查合格(1 ml/支);pH值为6.9;无菌试验合格;分装重量精密性为0.95%。结论制备的痘苗病毒标准品可作为痘病毒载体疫苗滴度测定用标准品。
Objective To prepare the national standard for determination of titers of virus vector-based vaccines.Methods Vaccinia virus was prepared with chick embryo fibroblasts（CEFs）,purified by density gradient ultracentrifuge,added with preservative for smallpox vaccine,then filled and stored in frozen,and distributed to four laboratories for cooperative calibration by plaque assay with crystal violet staining.The samples were diluted serially,and two dilutions were used for each test.Five parallel tests were performed on each dilution.The geometric mean of determination results in various laboratories was calculated,based on which the vaccinia virus titer in standard was defined.The virus standard after storage at various temperatures for various time durations were evaluated for stability,and subjected to overall control tests.Results A total of 44 tests were performed in four laboratories,and the results,with a geometric mean of 3.05 × 10^6 PFU / ml,a 95% CI of 2.00 × 10^6 ～ 4.65 × 10^6 PFU / ml,a standard deviation of 0.09 and a total CV value of 1.41%,showed no significant difference（P〉0.05）.The prepared vaccinia virus standard was stable after storage at 4℃ for 10 d,while was less stable at 37℃,and was qualified in filling quantity（1 ml per container） and sterility test,of which the pH value was 6.9 and the precision for filling was 0.95% based on weight.Conclusion The prepared vaccinia virus standard may be used for determination of titers of vaccinia virus vector-based vaccines.
出处
《中国生物制品学杂志》
CAS
CSCD
2012年第9期1099-1101,1105,共4页
Chinese Journal of Biologicals
基金
十一五"国家传染病科技重大专项(2009ZX10004-801)
关键词
痘苗病毒
蚀斑滴定
标准品
Recombinant vaccinia virus
Plaque assay
Standard
