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 continuous 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 intensification 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 x 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 x 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.
Jardin
Barbara Ann
BA
Biotechnology Research Institute, National Research Council Canada, 6100 Royalmount Avenue Montreal, Quebec, Canada H4P2R2.
Montes
Johnny
J
Lanthier
Stephane
S
Tran
Rosa
R
Elias
Cynthia
C
D000469:Alkaline Phosphatase / Q000096:biosynthesis* / Q000378:metabolism; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D019149:Bioreactors; D001709:Biotechnology / Q000379:methods*; D018929:Cell Culture Techniques / Q000379:methods; D049109:Cell Proliferation; D002478:Cells, Cultured; D010477:Perfusion; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D018411:Spodoptera
D011994:Recombinant Proteins; D000469:Alkaline Phosphatase