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目的:应用生物反应器培养Vero细胞和乙型脑炎病毒,大规模生产灭活乙脑疫苗。方法:采用乙型脑炎病毒株(京卫研P3株)构建毒种库,Vero细胞为培养基质,应用特定型号的生物反应器,投入25g/L的微载体,以灌流方式培养细胞,连续收获乙脑病毒液,经过浓缩、灭活、纯化等工艺制备病毒原液,并最终分装成疫苗制品。结果:细胞培养密度达到1.5*107个/毫升;病毒滴度8-9LogLD50/毫升;层析纯化的杂蛋白去除率大于99.9%,总蛋白含量小于10ug/剂,残余DNA含量小于10pg/剂。结论:应用生物反应器进行Vero细胞培养,病毒连续灌流收获,可大规模生产灭活人用乙型脑炎纯化疫苗。
Objective: To culture Vero cells and Japanese encephalitis virus using bioreactor and to produce large-scale inactivated JE vaccine. Methods: The virulence library was constructed by using Japanese encephalitis virus strain (Jing Wei-yan P3). The Vero cells were used as culture medium. The cells were cultured in perfusion mode by using a specific type of bioreactor and 25 g / L microcarrier. JE virus harvested, after concentration, inactivation, purification and other processes to prepare the virus stock solution, and finally divided into vaccine products. Results: The cell culture density reached 1.5 * 107 cells / ml; the virus titer was 8-9LogLD50 / ml; the chromatographic purification of the mixed protein was more than 99.9%, the total protein content was less than 10ug / dose and the residual DNA content was less than 10pg / dose. Conclusion: The bioreactor was used to culture Vero cells and the virus was continuously perfused and harvested for the mass production of inactivated encephalitis purified vaccine.