表达重组抗CD52单克隆抗体CHO细胞灌流培养基的筛选

Screening of perfusion culture medium for expressing recombinant anti-CD52 monoclonal antibody by CHO cells

目的筛选重组抗CD52单克隆抗体CHO细胞株培养和连续灌流表达用培养基,以提高抗体表达量。方法通过调整原有批培养用培养基中谷氨酰胺和植物水解蛋白,获得5种培养基配比。使用模拟灌注方式进行细胞培养,分析细胞密度、活细胞比率和目标蛋白表达,筛选连续灌流细胞培养和表达用培养基。最后在7 L反应器中采用灌注培养方式对筛选获得的培养基进行验证。结果使用50 mL细胞培养管进行模拟灌注培养时,活细胞比率较高,达到90%以上;CHO细胞在添加谷氨酰胺至4.0 mmol/L和植物水解蛋白至5.0 g/L的批培养用培养基中生长速度最快;在基础培养基中抗体表达量比优化前高15%。20 d培养周期内,优化的培养基在7 L反应器中可以维持CHO细胞密度在(2727±253)万个/mL,活细胞比率在95%以上。结论通过模拟灌注培养,筛选获得了一种在7 L反应器灌流培养中适宜于重组抗CD52单克隆抗体CHO细胞表达的培养基。

Objective To select the perfusion medium for culturing recombinant CHO cells expressing anti-CD52 monoclonal antibody in order to increase the target protein amount.Methods Five kinds of mediums were tested by adjusting the concentrations of glutamine and plant hydrolysate in the batch culture medium.The cell density,the ratio of living cells and the expression of target protein were analyzed by simulation perfusion,and a selected medium for continuous perfusion culture and expression was performed and verified in a 7 L bioreactor by perfusion culture.Results Cell viability was almost the same in different medium,cells divided fastest in the bath culture medium with additional glutamine to 4 mmol/L and plant hydrolyzed to 5.0 g/L.Antibody expression amount in the optimized medium was 15%higher than that of culture in the pre-optimization medium.Adjusted medium could maintain cell grow and protein expression in 7 L bioreactor within a 20 days of cultural cycle during this period,cell density was maintained at(27.27±2.53)×10^(6)/mL and cell viability was maintained above 95%.Conclusion A suitable medium for perfusion culture was obtained in a 7 L reactor for recombinant anti-CD52 monoclonal antibody with CHO cells by simulation perfusion culture.