高通量生物反应器Tubespin培养CHO细胞条件的优化  

Optimization of Condition for Culture of CHO Cells by Tubespin,a Disposable High-throughput Bioreactor

作　　者：谢奎[1] 雷云[1] 黄鹂[1] 王一婷[1] 熊盛[1] 

机构地区：[1]暨南大学生物医药研究开发基地广东省生物工程药物重点实验室基因药物国家工程中心,广州510632

出　　处：《中国生物制品学杂志》2011年第6期715-719,共5页Chinese Journal of Biologicals

基　　金：国家自然科学基金(30873082);广州市生物产业化重大专项(2010U1-E00541)

摘　　要：目的优化高通量生物反应器Tubespin培养CHO细胞的条件,提高蛋白表达量。方法采用Tubespin生物反应器在不同转速(160、180和200 r/min)和培养体积(5、10、20和35 ml)条件下振荡悬浮培养CHO细胞,测定不同条件下的气体交换速率、细胞密度、细胞活力及蛋白相对表达量,优化细胞培养条件。在最佳培养条件下,向培养基中分别添加30、60和90 mmol/L氯化钠,观察不同浓度氯化钠对细胞密度、细胞活力、摄氧率、溶氧及重组蛋白表达的影响。结果 Tubespin具有良好的气体交换速率,可满足高密度细胞培养的需要;经优化,细胞培养最佳条件为:摇床转速180 r/min,培养体积10 ml,此时细胞生长密度高,较快进入稳定期进行重组蛋白的表达。细胞培养72 h时向培养基中添加60 mmol/L氯化钠,能使重组蛋白的表达量提高1倍。结论优化了Tubespin培养CHO细胞的条件和氯化钠诱导浓度,提高了重组蛋白的产量,为大规模发酵生产奠定了基础。Objective To optimize the condition for culture of CHO cells by Tubespin,a disposable high-throughput bioreactor and increased the yield of protein.Methods CHO cells were subjected to suspension culture in oscillation by using Tubespin at various rotation speeds（160,180 and 200 r/min） in various volumes（5,10,20 and 35 ml）.The gas exchange rates,cell densities,cell viabilities and relative expression levels of protein under various conditions were compared,based on which the culture condition for cells was optimized.Under the optimal condition,the medium was added with 30,60 and 90 mmol/L sodium chloride respectively,and the effects on cell density,cell viability,oxygen uptake rate,dissolved oxygen and expression of recombinant protein were observed.Results Tubespin showed high gas exchange rate which met the necessity of high density cell culture.The optimal rotation speed shaker and culture volume of cells were 180 r/min and 10 ml respectively.Under the optimized condition,cells growth density was high,and the stable growth stage was reached rapidly,which was suitable for expression of recombinant protein.The addition of 60 mmol/L sodium chloride into medium 72 h after culture increased the expression level of recombinant protein by 1 fold.Conclusion The condition for culture of CHO cells by Tubespin as well as the concentration of sodium chloride for induction were optimized,and the yield of recombinant protein increased,which laid a foundation of large-scale production of recombinant protein by fermentation.

关 键 词：Tubespin CHO细胞 细胞培养技术 氯化钠 细胞密度 蛋白表达 

分 类 号：R392-33[医药卫生—免疫学]