大鲵虹彩病毒转瓶规模化培养条件研究

Condition Optimization for Large-Scale Cultivation of Chinese Giant Salamander Iridovirus by Rotary Bottle

研究了利用2.0 L转瓶进行大鲵虹彩病毒规模化培养的最佳工艺条件。结果显示,培养液稀释体积为150m L时,1×106个/m L的初始接种细胞密度可使大鲵虹彩病毒滴度达到最高;培养液稀释体积和接种细胞密度存在一定相关性,在不影响病毒滴度的情况下,可以在细胞密度为5×105个/m L时加入300 m L的培养(维持)液以尽可能多的收获病毒。在1∶20和1∶40的病毒接种剂量下,病毒均可大量增殖,滴度最高为104.35TCID50/0.1 m L;接毒方法简化步骤对病毒在转瓶中的增殖无明显影响。在转瓶中EPC细胞接种病毒的增殖动态曲线显示,接种病毒8 h后,病毒滴度开始上升并进入对数生长期,24 h后病毒增值速度趋缓,72 h病毒滴度达到最大。本研究为大鲵虹彩病毒的规模化培养和细胞培养疫苗的规模化制备技术奠定了基础。

The techniques for large-scale cultivation of Chinese giant salamander iridovirus（ CGSIV） were investigated and optimized by rotary bottle. Results showed that the optimal cell density was 1 × 106 ind. / m L when culture medium volume was 150 m L. Cell density was somewhat correlated to the culture medium volume,and 5 × 106 ind. / m L cell density as well as 300 m L culture media might harvest the most virus. When the inoculation dosage of seed virus was 1∶ 20 or 1∶ 40（ volume ratio of original virus liquid and culture medium volume）,the virus could be largely proliferated,and the viral titer might reach to 104. 35TCID50 /0. 1m L. No significantly negative effect on virus proliferation was observed from the simplified steps for rotary bottle. Proliferation dynamics of virus in EPC cells demonstrated that viral titer began to rise at the 8th h after inoculation,followed by a logarithmic increase,and gradually became gentle at the 24 thh,finally the viral titer reached its maximum at the 72 ndh after inoculation. The obtained data facilitated the large-scale preparation of vaccine against CGSIV.