Culture Process For Anti-EGFR Monoclonal Antibody

The culture technology of recombinant CHO-K1 cells with stable expression of monoclonal antibody against EGFR was studied.The basic culture medium(CDM4Per MAb)and cell lines were determined by Fed-batch culture on shaker and 3L reactor.Fed-batch culture was conducted on a 3L reactor to determine the effects of controlled culture temperature and p H on cell growth and protein expression.The results showed that:(1)when VCD≥8.0×10~6cells/ml or Day5 of the culture,the temperature dropped from 37.0℃to 33.0℃and the p H range was 7.00±0.2,the cells could maintain a high number of living cells,which were more suitable for cell growth and protein expression.The antibody expression was1863mg/L,but lactic acid began to accumulate on Day12,affecting the cell viability in the later period.The feeding strategy of Day10 and Day13 were changed into daily feed to effectively control the continuous accumulation of lactic acid,and the cell viability was kept above 80%.Cell adaptability and Top-off operation were tested on a 50L one-time WAVE reactor in order to simulate the seed chain amplified to 250L bioreactor.The results showed that:(1)the cells grew well in the WAVE reactor after the removal of medium pressure;(2)according to the passage data,it can be seen that the cells are passed on to the first generation in about 3 days,that is,it takes about 17 days for the culture cycle from flask resuscitation to n-1.The cell culture process was confirmed on the 50L SUB reactor,and the locked culture process was for the 250L one-time reactor.The confirmed process was Fed-batch cultured on SUB250L,and the antibody expression was 2117mg/L to complete the cell culture amplification process.