Effective Development of Intensified Perfusion Culture Based on Shake Tube Semi-Continuous Bioreactors and Optimal Osmolarity Constraint

The development of perfusion strategies supporting ultra-high cell growth and productivity for the intensified perfusion cultures (IPC) is very challenging. In this study, a novel approach was developed to address this issue by utilizing the shake tube semi-continuous bioreactors (ST) combining with a concentrated feed from the traditional fed-batch (TFB) platform. First, the impact of operating conditions in the ST were evaluated to identify those that best reflected the cell performance for the benchtop perfusion bioreactors (STR). Subsequently, the optimization of several key process parameters along with the metabolic data obtained for the design of the IPC was performed in the ST. Further, an empirical model to predict culture osmolarity was developed, verified, and subsequently utilized to determine the best combinations of the basal and feed perfusion rates ensuring the optimal osmolarity at the targeted viable cell density for the IPC process. Finally, the designed perfusion strategies were successfully validated in two IPC processes operated at 80 and 100×106 cells/m in the 3-L STR with an approximate 9-fold increase in cumulative titers compared to the TFB process. Overall, this study offers a novel and prospective method for the effective development of the IPC process to achieve significant process intensification.