Production of Recombinant Human Monoclonal Antibody Usingras-Amplified BHK-21 Cells in a Protein-free Medium

Abstract

A ras oncogene-amplified recombinant BHK-21 cell line (ras-rBHK-IgG) has been established, and was shown to hyperproduce the recombinant IgG chimeric human monoclonal antibody (hMAb) AE6F4, which recognizes lung cancer cells. We found that the ras-rBHK-IgG cell could be easily cultured in a protein-free ERDF medium supplemented with iron(III) nitrate, hydroxyethyliminodiacetic acid, and non-protein synthetic attachment factor as well as in a serum-free ERDF medium supplemented with insulin, transferrin, ethanolamine, and sodium selenite. The productivity of recombinant hMAb from the cells cultured in dishes at high cell densities was higher in protein-free medium than in serum-containing medium. True high density culture of the ras-rBHK-IgG cells was done in protein-free medium using the Tecnomouse, which is a novel hollow fiber bioreactor system. After culture for 30 days in protein-free culture, a total amount of about 14 mg of the recombinant hMAb AE6F4 was obtained, and was shown to be reactive against lung cancer cells in tissues.

Related effects of cell adaptation to serum-free conditions on murine EPO production and glycosylation by CHO cells

“…Indeed, serum contains many components (growth factors, insulin, transferrin, albumin, vitamins, etc....), which are known to be essential to several cellular functions like growth regulation or intracellular transports (Froud 1999). Thus, compared to serum-containing cultures, reduction of protein production and/or cell growth are generally observed when cells are transferred to serum-free media (Inoue et al 1996;Lee et al 1999;Ozturk et al 2003). Extensive investigations for optimising medium composition to a determined cell line cultured in the desired conditions are required to recover equivalent performances (Buntemeyer et al 1991;Hewlett 1991;Castro et al 1992;Liu et al 2001).…”

Improved gene transfer and normalized enzyme levels in primitive hematopoietic progenitors from patients with mucopolysaccharidosis type I using a bioreactor

“…The application of a hollow‐fiber bioreactor system (HBS) may provide a solution for many of the difficulties associated with hematopoietic stem cell gene transfer. In the past, such systems have been used for production of industrial and clinical‐grade reagents such as monoclonal antibodies 24, cell lines 25, 26, and other biologics destined for pharmacological applications 27, 28. In our previous work 29, we have demonstrated that an HBS could be used to culture and transduce human primary peripheral blood lymphocytes (PBL) in clinically useful quantities (up to 10 10 primary PBL).…”

Closed hollow-fiber bioreactor: a new approach to retroviral vector production

“…The initial studies of cell growth in a hollow‐fiber bioreactor demonstrated that solid tissue densities could be reached in vitro 8, 9 and that such culture could secrete 3‐ to 13‐fold more biologically active product than monolayer cultures10. Since then, such systems have been used for production of industrial and clinical‐grade reagents such as monoclonal antibodies11–13, cell lines14–16, and other biologics destined for pharmacological applications17, 18. Recently, an approach using the bioreactor system for culture and transduction of human primary lymphocytes has been reported to combine ultrafiltration and transduction in one step19.…”