CHO工程细胞(11G-S)悬浮培养的无血清培养基的设计

Serum-free medium for suspension culture of recombinant Chinese hamster ovary (11G-S) cells

以悬浮适应的表达重组尿激酶原(Pro-urokinase,pro-UK)CHO工程细胞系11G-S为对象,采用Plackett-Burman实验设计及响应面分析法,设计支持CHO工程细胞(11G-S)悬浮生长的无血清培养基。以细胞密度为评价指标,在单因素实验的基础上采用Plackett-Burman实验设计对影响细胞生长的培养基添加成分进行考察,确定了3种对细胞生长明显促进作用的培养基添加成分:胰岛素、转铁蛋白及腐胺。继而利用响应面法分析了这3种添加成分的最佳水平范围,设计了一种适用于CHO工程细胞(11G-S)悬浮培养的无血清培养基SFM-CHO-S。11G-S细胞在SFM-CHO-S批次悬浮培养的细胞最大生长密度达到4.12×106cells/mL,pro-UK的最大累积活性达到5614IU/mL,培养效果优于商品化的同类无血清培养基。

With suspension adapted recombinant Chinese hamster ovary （CHO） cell lines 11G-S expressing human pro-urokinase （pro-UK） as the object of study, a serum-free medium for the cultivation of recombinant CHO cells in suspension was formulated by using Plackett-Burman design and response surface methodology. The two-level Plackett-Burman design was used to evaluate the effect of 10 medium supplements on the growth of the llG-S cells in suspension culture. Among the 10 medium supplements, insulin, transferrin, and putrescine were identified as the most significant factors （P〈0.05）. The response surface methodology with three factors and three levels was used to determine the optimal levels of these factors. And a serum-free medium, SFM-CHO-S for recombinant CHO cells suspension culture was formulated. The maximum cell density of 11G-S cells in SFM-CHO-S in suspension batch culture reached 4.12× 10^6 cells/mL with a maximum pro-UK activity at 5 614 IU/mL, which was superior to the commercial serum-free medium for recombinant CHO cells.