中国仓鼠卵巢工程细胞无血清流加培养关键工艺参数的考察

Evaluation of the critical process parameters for the cultivation of recombinant Chinese hamster ovary cells in serum-free fed-batch mode

摘要:

主要考察流加培养基中不同营养成分、流加起始时间及初始接种密度对11G-S 细胞无血清流加培养的影响。在研究中以悬浮适应的表达尿激酶原 (Pro-urokinase，Pro-UK) CHO 工程细胞系11G-S 为研究对象，在100 mL 的摇瓶中无血清悬浮流加培养11G-S 细胞，同时以活细胞密度、细胞活力及Pro-UK 活性为评价依据。结果表明在培养基中氨基酸、无血清添加成分及无机盐对促进细胞生长、细胞活力维持及蛋白表达起着较为重要的作用；且流加起始时间为72 h 及初始接种密度为3×10^5<

Abstract:

Taking a suspension adapted recombinant CHO cell line, 11G-S expressing human Pro-urokinase (Pro-UK) as the object of study, the impacts of different feeding nutrients, the start time of feeding and cell inoculation density on the growth and Pro-UK production of 11G-S cells in serum-free fed-batch culture were evaluated in 100 mL shacking flasks. The results indicated that amino acids, serum-free supplements and inorganic salts played important role in cell growth, cell viability and protein expression. And the effects of cells fed-batch culture was much better with the initial cell inoculation density at 3×10⁵~4×10⁵ cells/mL and the start time of feeding set at 72 h, a maximum viable cells density of 7.8×10⁶ cells/mL with a peak Pro-UK activity at 8 570 IU/mL was achieved during 12 d fed-batch culture. Further, the μ of the 11G-S cells at the middle phase of the fed-batch culture, and both the q_glu and q_gln of the 11G-S cells at the middle and later phases of the fed-batch culture was higher than that of the 11G-S cells at the same phase of the batch culture, respectively.