Controlling Apoptosis to Optimize Yields of Proteins from Mammalian Cells
Apoptosis is the foremost method of cell death in bioreactors and can be caused by nutrient limitation, toxin accumulation, and growth factor withdrawal. By delaying the onset of this form of programmed cell death, one can achieve longer sustained viabilities in culture, thereby increasing product yield. Described here is a genetic-based, step-by-step method to generate an apoptosis-resistant cell line. This cell line, then, can be used as a platform for biotherapeutic protein production. The key steps include antiapoptotic transgene selection and transfection followed by clonal isolation and screening. With the proper screening methods, one can obtain a robust cell line that resists the harsh conditions of late-stage and/or high-density culture.
Key wordsApoptosis Transfection Selection Screening Flow cytometry bcl-XL CHO Monoclonal antibody Mammalian cell culture
- 1.DePalma, A. (2008) Strengthening mammalian cell culture. Genetic Engineering & Biotechnol. News. 28.Google Scholar
- 4.Nivitchanyong, T., Martinez, A., Ishaque, A., Murphy, J.E., Konstantinov, K., Betenbaugh, M., and Thrift, J. (2007) Anti-apoptotic genes Aven and E1B-19 K enhance performance of BHK cells engineered to express recombinant factor VIII in batch and low perfusion cell culture. Biotechnol. Bioeng. 98, 825–841.PubMedCrossRefGoogle Scholar
- 6.Dorai, H., Nemeth, J., Cammaart, E., Wang, Y., Tang, Q., Magill, A., Lewis, M.J., Raju, T.S., Picha, K., O’Neil, K., Ganguly, S., and Moore, G. (2009) Development of mammalian production cell lines expressing CNTO736, a glucagon like peptide-1-MIMETIBODYTM: Factors that influence productivity and product quality. Biotechnol. Bioeng. 103, 162–176.Google Scholar
- 8.Ishaque, A., and M. Al-Rubeai. (2007). Measurement of apoptosis in cell culture, in Methods in Biotechnology, Animal Cell Biotechnology (Ralf Portner, ed.) Human Press, Totowa.Google Scholar
- 9.Figueroa Jr., B., Ailor, E., Osborne, S., Hardwick, J.M., Reff, M., and Betenbaugh, M.J. (2006) Enhanced cell culture performance using inducible anti-apoptotic genes E1B-19 K and Aven in the production of a monoclonal antibody with Chinese hamster ovary cells. Biotechnol. Bioeng. 97, 877–892.CrossRefGoogle Scholar