Vero细胞无血清培养基的筛选及其培养流感病毒H5N1条件的优化  

Screening of serum-free medium for Vero cells and potinization of condition for culture of influenza H5N1 virus

作　　者：李在晗 苏杨起 常亚军[1] 杨家方 王婷婷[1] 杨耀云[1] 刘圆圆[1] 杨建波 唐聪[1] 徐丽兰[1] 张恒[1] 廖国阳[1] 李卫东[1] 

机构地区：[1]中国医学科学院北京协和医学院医学生物学研究所,云南昆明650118 [2]南开大学生命科学学院,天津300071

出　　处：《中国生物制品学杂志》2016年第4期407-412,共6页Chinese Journal of Biologicals

基　　金：细胞流感减毒活疫苗合作研究(国际科技合作项目;2011DFR30420);腮腺炎减毒活疫苗等病毒性疫苗关键技术及产品研发(2012AA02A404);Vero细胞流感减毒活疫苗研制(2013ZX10004003003002);疫苗研发与新技术应用省创新团队(2015HC027)

摘　　要：目的筛选适宜Vero细胞培养的无血清培养基,并优化其培养流感病毒H5N1的条件。方法取VP-SFM培养的Vero细胞(SFM-Vero),以增殖速率为指标,筛选无血清培养基[VP-SFM、EX-CELL^(TM) VERO、Virus Pro ^(TM) VERO、自制无血清培养基1(SFM-1)和2(SFM-2)];采用中心组合实验(Box-Behnken)优化SFM-Vero培养流感病毒H5N1的pH、TPCK-Trypsin浓度和病毒接种MOI。结果 Vero细胞在VP-SFM和自制SFM-1培养基中生长情况好于其他无血清培养基;H5N1病毒株能够感染无血清适应的Vero细胞;SFM-Vero培养流感病毒H5N1较优条件为:p H 7.64,TPCK-Trypsin浓度0.68μg/ml,MOI 0.01。在该条件下培养的流感病毒HA滴度可达768 HAU/50μl,为正常培养基培养的Vero细胞的2.33倍。结论筛选出适宜Vero细胞生长的无血清培养基,优化了流感病毒H5N1 Vero细胞适应株无血清培养的条件。Objective To screen a serum-free medium（SFM） for Vero cells and optimize the condition for culture of influenza H5N1 virus the medium. Methods The Vero cells were cultured in VP-SFM, EX-CELLTMVERO, Virus ProTM VERO and self-made serum free medium 1（SFM-1）and 2（SFM-2）, and determined for multiplication rate to screen SFM.Box-Behnken experiment design was applied to optimize the p H value, working concentration of TPCK-Trypsin and MOI for inoculation of culture of influenza H5N1 virus in SFM-Vero cells. Results The growth situations of Vero cells in VPSFM and SFM-1 were better than those in other SFM. Serum-free adapted Vero cells were infected with influenza H5N1 virus. The optimal p H value, working concentration of TPCK-Trypsin and MOI for culture of influenza H5N1 virus in SFMVero cells were 7. 64, 0. 68 μg / ml and 0. 01 respectively. The HA titer of influenza H5N1 virus cultured under the optimal condition was 768 HAU / 50 μl, which was 2. 33 times of that cultured in normal medium. Conclusion The SFM suitable for Vero cells was screened, and the condition for serum-free culture of Vero cells adapted strain of influenza H5N1 virus was optimized.

关 键 词：无血清培养基 流感病毒Vero细胞适应株 H5N1 

分 类 号：R373.1[医药卫生—病原生物学]