转瓶培养与生物反应器微载体培养乙脑病毒的比较

分别用15L转瓶与15L生物反应器微载体(2.5g/L CytodexⅢ)系统培养Vero细胞并接种乙型脑炎病毒(简称乙脑病毒)。转瓶培养Vero细胞7~8d,细胞数最高能达到8×108;当单层细胞长至3.0~4.5×108时接种乙脑病毒,病毒滴度能达到6.5~6.98 lg PFU/ml,并能够连续收获4~5次;采用微载体系统培养Vero细胞,细胞密度最高能达到170×108;当单层细胞长至60~70×108时接种乙脑病毒,病毒滴度能达到7~7.5 lg PFU/ml,并能够连续收获13~15次。两种方式培养的乙脑病毒收获液分别经灭活、浓缩、柱层析纯化后制备Vero细胞乙脑纯化疫苗,各项检定指标均符合《中国药典》的相关要求。

Comparisons of the cell cultivation between using microcarrier in a 15L bioreactor and roller bottle for the preparation of JEV vaccine

Japanese encephalitis virus(JEV) were separately cultivated in Vero cells with 15L roller bottle and 15L bioreactor system(2.5g/L microcarrier、Cytodex Ⅲ).A higher cells density was recovered from cultures using 15Lbioreactor(170×108 cells),compared with using the roller bottle mode(8×108 cells).JEV were innoculated into vero cells when the cell yields to 60-70×108 in the bioreactor and 3-4.5×108 in roller bottle,respectively.After 3-4 days the virus titres reached to 7-7.5lgPFU/ml and the harvest times were 13-15 for the bioreactor while the virus titres were 6.5-6.98 lg PFU/ml and the harvest times were 4-5 for the roller bottle.The JEV vaccine(final bulk)were prepared separately using the harvested virus by inactivation,concentration and purification and the results detected could comply with the requirements of Chinese pharmacopoeia.