麻疹减毒活疫苗细胞工厂多次收获工艺探索

Exploration of measles live attenuated vaccine multiple harvest by cell factory

目的建立麻疹减毒活疫苗细胞工厂多次收获工艺。方法采用细胞工厂培养2BS细胞,以不同感染复数接种麻疹病毒,绘制病毒生长曲线以确定适宜感染复数。在感染后不同时间点,微量滴定法比较仅收获胞外病毒与收获胞内外病毒的滴度。比较单次收获与多次收获病毒的滴度,并对原液和冻干成品进行稳定性试验。结果确定细胞工厂培养麻疹病毒的适宜感染复数为0.010~0.100。感染后108~168 h细胞外病毒滴度与含胞内病毒滴度无明显差异(t=0.524,P>0.05)。多次收获得到的病毒总量高于单次收获。单次收获与多次收获得到的病毒原液-65 ℃保存3个月,病毒滴度下降均不超过每毫升0.3 lg半数细胞物培养感染量,冻干成品37 ℃保存1周,滴度下降均不超过每毫升0.5 lg半数细胞培养物感染量。结论建立了细胞工厂培养麻疹减毒活疫苗的多次收获工艺。

Object To establish a multiple harvest cell factory production process of measles live attenuated vaccine.Methods 2BS cells were cultured in cell factory and inoculated with different multiplicities of infection(MOIs).Growth curve of the virus was drawn to determine a suitible MOI.At different time points after infection,titers of extracellular virus and total virus were tested by microplate titration.Titers of virus by single harvest and multiple harvest were compared,and stability of virus bulks and final products by two methods was tested.Result The suitable MOI of measles virus in cell factory culture was 0.01-0.10.There was no significant difference(t=0.524,p>0.05)on virus titers between the extracellular virus and total virus at 108-168 h after infection.The total amount of virus obtained by multiple harvesting was higher than that of single harvesting.Titers of virus bulks by two methods after being stored at-65℃for 3 months dropt by less than 0.3 lg 50%cell culture infectious dose(CCID50)per ml,and titer of final products by two methods after being stored at 37℃for 1 week dropt by less than 0.5 lgCCID50/ml.Conclusion The multiple harvest cell factory production process of measles live attenuated vaccine is established.