Vero细胞微载体培养的化学成分明确无血清培养基的设计

目的：设计适用于Vero细胞微载体培养的化学成分明确无血清培养基。方法：以商品化的DMEM／F12合成培养基为基础培养基，应用Plackett—Burman实验设计和响应面分析法设计支持Vero细胞微载体培养的化学成分明确无血清培养基。结果：以细胞密度为评价指标，在单因素实验的基础上采用Plackett-Burman实验设计考察10种培养基添加成分对Vero细胞生长的影响，确定了3种对Vero细胞生长起明显促进作用的培养基添加成分，为胰岛素、血清素和腐胺。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，设计了一种支持Vero细胞贴附培养的无血清培养基（VERO—SFM—A）。在Bellco搅拌式培养瓶中采用VERO-SFM．A和Cytodex1微载体培养Vero细胞，细胞密度由接种时的4×10^5cells／ml增加到培养6d后的22．3×10^cells／ml，细胞活力保持在96％以上。结论：VERO—SFM—A能够有效地支持Vero细胞在微载体表面固定化生长并达到较高的细胞密度，具有实际应用于Vero细胞微载体规模化培养的应用潜力。

Development of a Chemically Defined Serum-Free Medium for Vero Cells Grown on Microcarriers

Objective： To develop a chemically def＇med serum-free medium for Vero cells grown on microcarriers. Methods： With the commercially synthesized medium DMEM/F12（1：1, v/v） as the basal medium, the effect often medium supplements on the growth of Vero cells in anchorage culture mode was evaluate by using Plackett-Burman design and response surface methodology. Results： Among the ten medium supplements tested, insulin, serotonin and putrescine were identified as the most significant factors （P〈0.05） for promot- ing the growth of Vero cells. The optimal levels and mutual-influence of insulin, serotonin and putrescine for the growth of Vero cells was 3.5 mg/L, 32 μmol/L and 7 mg/L, respectively. And a chemically defined serum-free medium designated VERO-SFM-A for Vero cells grown on microcarriers was formulated. With an inoculated density of 4 × 10^5 cells/ml in VERO-SFM-A, the Vero cells grown on Cytodex-1 in Bellco spinner flask reached the density of 22.3 × 10^5 cells/ml with the cell viability maintained above 96% after six-day cultivation. Conclusion： VERO-SFM-A could support the growth of Vero cells immobilized on microcarriers with a satisfy cell density, which indicated the ootential ofbeing used in large scale Vero cells culture on microcarriers.