High cell density fed batch and perfusion processes for stable non‐viral expression of secreted alkaline phosphatase (SEAP) using insect cells: Comparison to a batch Sf‐9‐BEV system

The development of insect cells expressing recombinant proteins in a stable continuous manner is an attractive alternative to the BEV system for recombinant protein production. High cell density fed batch and con- tinuous perfusion processes can be designed to maximize the productivity of stably transformed cells. A cell line (Sf-9SEAP) expressing high levels of the reporter protein SEAP stably was obtained by lipid-mediated transfection of Sf-9 insect cells and further selection and screening. The expression of the Sf-9SEAP cells was compared with the BEVS system. It was observed that, the yield obtained in BEVS was similar to the batch Sf-9SEAP at 8 and 7 IU/mL, respectively. The productivity of this foreign gene product with the stable cells was enhanced by bioprocess intensifica- tion employing the fed-batch and perfusion modes of culture to increase the cell density in culture. The fed batch process yielded a maximum cell density of 28 � 10 6 cells/mL and 12 IU/mL of SEAP. Further improvements in the productivity could be made using the perfusion process, which demonstrated a stable production rate for extended periods of time. The process was maintained for 43 days, with a steady-state cell density of 17-20 � 10 6 cells/mL and 7 IU/mL SEAP. The total yield obtained in the perfusion process (394 IU) was approximately 22 and 8 times higher than that obtained in a batch (17.6 IU) and fed batch (46.1 IU) process, respectively. Biotechnol. Bioeng. 2007;97: 332-345. 2006 Wiley Periodicals, Inc.