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

摘要:

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4.12×106 cells/mL，pro-UK的最大累积活性达到5 614 IU/mL，培养效果优于商品化的同类无血清培养基。

Abstract:

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 11G-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 4.12×106 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.