5L生物反应器中长期灌流培养CHO工程细胞生产rt-PA  

Production of rt-PA by long term cultivation of recombinant CHO cells in 5L bioreactor

作　　者：林福玉[1] 陈昭烈[1] 刘红[1] 吴本传[1] 李世崇[1] 黄培堂[1] 

机构地区：[1]军事医学科学院生物工程研究所,北京100071

出　　处：《军事医学科学院院刊》2000年第1期44-48,共5页Bulletin of the Academy of Military Medical Sciences

基　　金：国家"八六三"高技术基金!资助课题 (Z1 8 0 3 1 2 )

摘　　要：目的 :利用 5L反应器培养CHO工程细胞生产重组组织型纤溶酶原激活剂 (recombinanttissueplasminogenactivator ,rt PA)。 方法 :在 5L搅拌式生物反应器中采用Cytopore1多孔微载体和无血清培基DF5S连续灌流培养CHO工程细胞株 4B3。结果 :持续培养达 10 3d ,活细胞密度和rt PA生产水平分别达到 6 .52× 10 6 个细胞 /ml和 1716 1U/ml。细胞培养上清经StreamlineSP离子交换层析和Lysine Sepharose 4B亲和层析两步纯化 ,获得纯度达到 98%的rt PA。SDS PAGE分析表明 ,整个培养过程所生产的rt PA具有较好的质量一致性。产品热原检测合格。结论 :实现高密度长期培养 4B3细胞 ,确定 5L培养工艺 。Objective:To produce the CHO derived recombinant t PA in 5L bioreactor. Methods:Continuous perfusion cultivation of recombinant CHO cell line(4B3) cells was performed in a 5 L stirred bioreactor with the serum free medium DF5S and Cytopore1 as carriers. Result: The culture term was 103 days. The highest viable cell density of 4B3 and the highest production level of rt PA was 6.52×10 6 cells/ml and 17 161U/ml, respectively. Culture medium of 4B3 cells was purified with streamline SP ion exchange chromatography and Lysine Sepharose 4B affinity chromatography, the purity of rt PA reached 98%. Analysis by SDS PAGE showed that the quality of rt PA was consistent during the whole period of cultivation. Conclusion:Long term ,high density 4B3 cell culture in 5 L bioreactor was performed. It lays a foundation for large scale cell culturing for rt PA.

关 键 词：细胞培养 CHO工程细胞 生物反应器 RT-PA 

分 类 号：R318.0[医药卫生—生物医学工程]