Process design and development of a mammalian cell perfusion culture in shake‐tube and benchtop bioreactors

Abstract The development of mammalian cell perfusion cultures is still laborious and complex to perform due to the limited availability of scale‐down models and limited knowledge of time‐ and cost‐effective procedures. The maximum achievable viable cell density (VCD max ), minimum cell‐specific perfusion rate (CSPR min ), cellular growth characteristics, and resulting bleed rate at steady‐state operation are key variables for the effective development of perfusion cultures. In this study, we developed a stepwise procedure to use shake tubes (ST) in combination with benchtop (BR) bioreactors for the design of a mammalian cell perfusion culture at high productivity (23 pg·cell −1 ·day −1 ) and low product loss in the bleed (around 10%) for a given expression system. In a first experiment, we investigated peak VCDs in STs by the daily discontinuous medium exchange of 1 reactor volume (RV) without additional bleeding. Based on this knowledge, we performed steady‐state cultures in the ST system using a working volume of 10 ml. The evaluation of the steady‐state cultures allowed performing a perfusion bioreactor run at 20 × 10 6 cells/ml at a perfusion rate of 1 RV/day. Constant cellular environment and metabolism resulted in stable product quality patterns. This study presents a promising strategy for the effective design and development of perfusion cultures for a given expression system and underlines the potential of the ST system as a valuable scale‐down tool for perfusion cultures.