文献类型：期刊文献

中文题名：无血清培养MDCK细胞生产狂犬病病毒条件的初步优化

英文题名：Preliminary Optimization of Production Condition of Rabies Virus with MDCK Cells Cultured in Serum-free Medium

作者：樊庆帅[1];牛红星[1];吴书军[2];张旭[1];谭文松[1]

机构：[1]华东理工大学生物反应器工程国家重点实验室,上海200237;[2]上海生物制品研究所第二研究室,上海200052

年份：2009

卷号：22

期号：11

起止页码：1136

中文期刊名：中国生物制品学杂志

外文期刊名：Chinese Journal of Biologicals

收录：CSTPCD;;Scopus;CSCD:【CSCD2011_2012】；

语种：中文

中文关键词：MDCK细胞;无血清培养基;微载体培养;狂犬病病毒

外文关键词：MDCK cells; Serum-free medium; Microcarrier culture; Rabies virus;

摘要：目的开发一种适应于MDCK细胞生长和狂犬病病毒生产的无血清培养基MDCK-SFM,并对MDCK细胞生产狂犬病病毒的条件进行初步优化。方法方瓶培养MDCK细胞,考察混合脂类、微量元素、氢化可的松、酵母抽提物和鱼胨水解物对MDCK细胞生长的影响。转瓶微载体批培养MDCK细胞,以获得细胞在不同培养基中的生长动力学。以不同的感染量(MOI)和感染时间(TOI)接种狂犬病病毒,确定MDCK细胞感染狂犬病病毒的最佳MOI与TOI。比较含10%NCS的DMEM、VP-SFM和MDCK-SFM培养基培养MDCK细胞生产狂犬病病毒的效果。结果混合脂类有利于细胞贴壁,酵母抽提物和鱼胨水解物可不同程度地促进MDCK细胞生长。以MDCK-SFM培养基培养的MDCK细胞最高密度可达14.3×105个/ml。在MOI=0.5、TOI=72h的条件下,可得到最高的病毒滴度。MDCK-SFM培养基培养MDCK细胞生产狂犬病病毒,可获得较高的病毒滴度(5.13lgFFU/ml)。结论MDCK-SFM培养基适于MDCK细胞生长和培养狂犬病病毒,且成本低,成分比较明确,在MDCK细胞高密度培养生产狂犬病疫苗方面具有较高的应用价值。
Objective To develop a serum-free medium for culture of MDCK cells and production of rabies virus, and preliminarily optimize the production condition of rabies virus with MDCK cells. Methods MDCK ceils were cultured in a flask, and the effects of lipid mixture, trace element, hydrocortisone, yeast extract and fish peptone hydrolysate on cell growth were investigated. MDCK cells were cultured in microcarriers in spinner to obtain the dynamic curves of cell growths in various media. Rabies virus was in- oculated to MDCK cells at various MOls and TOls to optimize the condition of virus infection. The results of production of rabies virus with MDCK cells cultured in DMEM containing 10% NCS, VP-SFM and MDCK-SFM media were compared. Results Lipid mixture was helpful to cell adherence, and yeast extract and fish peptone hydrolysate promoted the growth of MDCK cells at different degrees. The highest density of MDCK cells cultured in MDCK-SFM medium reached 14. 3 × 10^5 cells/m/. The optimal MOI and TOI for infection of MDCK cells with rabies virus were 0. 5 and 72 h respectively. High virus titer （5. 13 lg FFU/ml） was obtained by culture of MDCK cells in MDCK-SFM medium. Conclusion MDCK-SFM medium was suitable for growth of MDCK cells and the culture of rabies virus. With low cost and definite component, the medium was of high value in production of rabies vaccine by high density culture of rabies virus in MDCK cells.