CHO Cell Culture Process And Metabolic Analysis In Tubespin

A novel high-throughput bioreactor, Tubespin was applied for cell culture and its competitive advantage than spinner flask was demonstrated. Then culture conditions in Tubespin for CHO cells were optimized as well as metabolic analysis to improve the cell concentration and the productivity of recombinant protein.CHO cells were cultured in tubespin and spinner respectively, cell density, viability, dissolved oxygen, productivity of re-protein and PH value were made into comparison. At last, the volume gas transfer values of the two bioreactors were measured. The results showed that cell cultured in tubespin was better than in spinner. The highest cell density of CHO DG44 and CHO TNF were 6.0×105 cells·ml-1 and 8.5×105 cells·ml-1 respectively, the dissolved oxygen in medium always above 80% air-saturation, PH value always above 6.7; but the highest cell density of the two cells in spinner were both under 4×105 cells·ml-1, the dissolved oxygen in medium declined to 1.9% and 0% respectively, PH value at last declined to 6.2, and more important, the productivity of recombinant protein when cell cultured in spinner was less than in tubespin. The volume gas transfer value of tubespin as high as 21.5 h-1, but it was only 2.7 h-1 of spinner. Then the culture conditions in tubespin were optimized, the cell density and protein productivity were both optimum at shaking speed 180 rpm with 10 ml working volume. The best concentration of sodium chloride which promote protein expression was optimized at this optimum condition, when the concentration of sodium chloride was 60 mM, the productivity of recombinant protein doubled.DMEM/F12 was selected as the basal medium through experimentation for the serum-free medium optimization. The optimum concentration of EAA, NEAA, VITAMINS, LIPIDS was studied by orthogonal test, Chemical defined medium sfml was gained. Then different peptone was added and sfm2 was gained. The results of cell culture and protein expression showed that sfm2 is better than commercialized medium CD DG44, near to P8 medium, and its price is only a half to them.The main metabolic parameter of CHO cell were analysised, it was found that the rate of consumption of glucose up to 80%; among the 17 amino acids, cystine has the most consumption which up to 100%, its used up may made a nutrition limited; During the culture, the concentration of lactate increased as well as it of ammonia, but both of them did not reach to the critical density which will inhibit cell growth; The variability of dissolved oxygen concentration was in parall with cell density, when cell density increase, it decrease; OUR was in parall with the phase of cell, at lag phase, OUR increased gradually, and up to the highest at log phase.