Study On Cell Culture Technology Of Recombinant Bevacizumab

Bevacizumab is a monoclonal antibody,which plays an important role in the treatment of tumor and macular degeneration.This research group has been built and select a stable expression of beacizumab bead sheet resistance of recombinant Chinese hamster ovary?recombinant Chinese hamster ovary,r CHO?cells,this article plans to the cell lines,cell culture technology was studied,and the cell line problems existing in the training process are studied and discussed.At first,by single factor experiment preliminary screening for five culture parameters of recombinant CHO cells,including the initial viable cell density?IVCD?,basic culture medium,culture temperature,shift viable cell density?SVCD?and the feed medium,the results showed that lactic acid accumulation in basic medium,thus further by adjusting the feeding method and adding copper ions on lactic acid metabolism regulation.Finally,it was determined that when the copper ion concentration was 120?g/L and the glucose concentration was 2 g/L on the second day from the 3rd day to the 14th day of cell culture,it was most beneficial to the cell growth metabolism and the expression of antibody products.In addition,the cell culture process of recombinant bevacizumab was optimized by experimental design?full factor design and center composite design?and response surface analysis,and the optimal process parameters were verified.The optimal combination is the mixture ratio of basal medium?BM04 ratio 75%?,Cu²⁺addition concentration?140?g/L?,the density of living cells in initial culture?0.9×10⁶ cells/m L?,the density of living cells in variable temperature?20×10⁶ cells/m L?and the culture time?14 days?.Besides,the method of detecting the heterogeneity of antibody charge and antibody concentration was validated.At the end of this study,the effects of metal ions(Cu²⁺/Mn²⁺)and fucose on the glycosylation of the cell antibody were preliminarily explored.The results showed that copper ion and manganese ion had a synergistic effect on the synthesis of G0 and afucosylated glycosylation of cell antibodies,but the high concentration of copper ion inhibited the production of antibodies,resulting in the decrease of antibody concentration.In this paper,the cell culture process of recombinant bevacizumab was preliminarily studied.The lactic acid metabolism was regulated by adjusting the feeding method and the concentration of copper ions,and the tolerance of cell culture was improved.Although the research on cell antibody glycosylation is not in-depth enough,the conclusions obtained in this paper have important reference value for the development or research of similar cell flask shaking culture process,and also provide research ideas and fundamental data for reactor process amplification and large-scale production.