【摘要】 目的探讨狂犬病病毒(Rabies virus,RV)aG株在原代鸡胚成纤维细胞上的传代适应性。方法将RV北京株固定毒10aG株豚鼠脑毒种在鸡胚成纤维细胞上连续传代培养,显微镜观察细胞的形态变化,ELISA检测各代次收获液中的病毒量,取11、21、31代病毒收获液,小鼠脑内毒力测定法检测病毒滴度。利用正交试验筛选最适病毒培养条件,以筛选的最佳比例接种病毒,绘制病毒生长曲线,并进行特异性及免疫原性检测。结果狂犬病病毒aG株感染鸡胚成纤维细胞未观察到明显的细胞病变效应,随着传代次数的增加,病毒含量呈上升趋势,11、21、31代病毒收获液的滴度分别为4.0、7.7和7.17 LogLD50/ml;多次传代后,最适培养条件为细胞浓度150×104个/ml、接种比例1∶500、血清浓度2%、收获时间为5 d;病毒连续收获3次后,细胞开始脱落,经检测其病毒滴度可达7.0 logLD50/ml以上,中和指数可达8 500,第21代病毒制备成的试制疫苗效价为2.0 IU/ml。结论 RV北京株固定毒10 aG株豚鼠脑毒种可适应于鸡胚细胞培养,为鸡胚细胞狂犬病疫苗的研究奠定了基础。更多还原

【Abstract】 Objective To investigate the adaptability of rabies virus(RV) aG strain in primary chicken embryo fibroblasts.Methods The fixed RV 10aG strain cultured in guinea pig brain was subcultured continuously in chicken embryo fibroblasts.The morphology of chicken embryo fibroblasts was observed by microscopy,and the culture condition for RV was optimized by orthogonal test.RV was inoculated at optimal MOI,of which the growth curve was plotted,and the specificity and immunogenicity were determined.Results No obvious CPE was observed in chicken embryo fibroblasts infected with RV aG strain.The RV content showed an increasing tendency with the increased passages.The titers of virus harvests of passages 11,21 and 31 were 4.0,7.7 and 7.17 LogLD50 / ml respectively.The optimal fibroblast concentration,MOI,serum concentration and time for harvest for culture of RV after subculture for several passages were 150 × 104 fibroblasts / ml,1 ∶ 500,2% and 5 d respectively.Shedding of the fibroblasts were observed after the virus was harvested for 3 times.The titer and neutralizing index of RV were more than 7.0 logLD50 / ml and 8 500 respectively.The potency of vaccine prepared with the RV of passage 21 was 2.0 IU / ml.Conclusion Fixed RV 10aG strain cultured in guinea pig brain may be adapted in chicken embryo fibroblasts,which lays a foundation of preparation of rabies vaccine in chicken embryo fibroblasts.更多还原