抗CD20抗体高产细胞株的筛选及质量评估

目的:筛选高表达单克隆细胞株,并通过优化培养基及流加物,最终达到提高目的蛋白产量及质量的目的。方法:通过有限稀释法对转染目的蛋白的CHO-S细胞进行单克隆化,应用双抗夹心ELISA方法对单克隆细胞株抗体表达量进行初步评估,最后根据筛选细胞株的活率、密度、产量及代谢情况,选择2~3株单克隆细胞进行培养条件优化,并对获得的发酵液进行纯化捕获,根据抗体蛋白表达量、糖型、等电点、纯度、酸碱峰分布等进行相应的评估分析,筛选出最优细胞株及最优培养方案。结果:经过单克隆化处理以及培养条件优化,蛋白的表达量由初始的不到500mg/L提升到2 290mg/L,且抗体蛋白纯度高达97.48%。抗体蛋白质量分析结果显示B1方案为该实验最优培养方案。结论:通过细胞株筛选、培养基优化能显著提高抗体蛋白的产量及质量,同时对抗体蛋白糖型、等电点、纯度等均有一定程度的优化。因此工业生产中可以通过高表达克隆的筛选、培养工艺优化等对目的蛋白产量及质量进行一定程度的改善与提高,对后期实验研究及工业化方案开发都具有很好的指导意义。

Anti-CD20rh MAb Quality Evaluation and Monoclonal Cell Line Screening

Objective: To screen high expression monoclonal cell lines and optimize the production and quality of Anti-CD20rh MAb. Method: Screen high expression monoclonal cell lines by finite dilution method, and evaluated the yield of monoclonal cell lines by double-sandwich ELISA. In order to screen the best cell culture program, two to three cells were picked based on the cells’ growth state, yield, viability, and so on. The result were evaluated by the sugar type, the isoelectric point, the purity and the distribution of the acid and alkali base peak. Results: The production of CHOS cells tripled after a series of optimization, increased from nearly 500mg/L to 2 290mg/L. After the optimization of culture program, the purity of the target protein reached up to 97.48%, and the distribution of the acid and alkali base peak looks more close to the ideal state. Conclusion: The production and quality of the target antibody were optimized by cloning and culture optimization, these results are of great significance to the later experimental research and industrial production.