Biphasic addition strategy of hypoxanthine and thymidine for improving monoclonal antibody production

In our previous study, we demonstrated that combinatorial addition of hypoxanthine (10 mg/L) and thymidine (2 mg/L) was able to stimulate initial cell growth and elevate volumetric concentration of antibody by 22% (Chen et al., Appl. Microbiol. Biotechnol., 93, 169–178, 2012). In this study, a systematic study was carried out to investigate the effects of hypoxanthine and thymidine (H&T) on cell growth and antibody production in a much wider range of concentration. In addition, we pursued to establish a highly productive fed-batch culture via rationally designing H&T addition regime. It was found that both cell growth and antibody production in batch cultures were H&T concentration-dependent. Specifically, a low concentration stimulated cell growth while exerting no influence on specific productivity (qmAb), and a high concentration inhibited cell growth, however, significantly enhancing qmAb. Subsequent experiments with fed-batch shaking flasks demonstrated the feasibility of improving antibody production using a biphasic addition strategy for H&T: supplementing a low concentration of H&T during initial cell growth phase and a high concentration of H&T at the production phase. By applying the optimized feeding regime, a maximum viable cell density (VCD) of 6.45 × 106 cells/mL and volumetric antibody production of 632 mg/L were achieved in a 2 L-B.Braun bioreactor. Taken together, in this study, a biphasic H&T addition strategy for cell culture was developed, which hold great promise to improve antibody production.