Abstract
Thirteen vitamins, twenty amino acids, hormones, inorganic salts, and other chemical agents, which constitute typical serum-free media, were evaluated for the development of fortified medium to enhance cell growth and productivity of recombinant antibody in the cultures of the recombinant Chinese hamster ovary (rCHO) cells. Two different rCHO cell lines, rCHO-A producing recombinant antibodies against the human platelet and rCHO-B secreting recombinant antibodies against the S surface antigen of Hepatitis B, respectively, were cultivated in batch suspension mode. Concentration of interested component in the tested medium was doubled to examine the fortification effect. Growth of rCHO-A cell and its antibody production were slightly improved with addition of either choline chloride, folic acid, thiamine⋅HCl, or LongTMR3IGF-I. On the other hand, in the cultivation of rCHO-B cell which was more sensitive to its environmental changes, hormones such as LongTMR3IGF-I and triiodothyronine (T3) as well as various vitamins involving choline chloride, i-inositol, niacinamide, pyridoxine HCl, and thiamine⋅HCl enhanced the cell growth and antibody production. Particularly, when concentration of consuming amino acid was doubled, remarkable increase in specific productivity was served, resulting in high final antibody concentration. These results were believed to provide a fundamental strategy of medium fortification useful for improvement of recombinant antibody production in serum-free medium.
Keywords: Chinese hamster ovary cell, Medium fortification, Protein- free medium, Recombinant antibody production, Serum-free medium
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Footnotes
These authors contributed equally to this work
Contributor Information
Ho Nam Chang, Phone: +82-42-869-3912, FAX: +82-42-869-8800, Email: hnchang@mail.kaist.ac.kr.
Duk Jae Oh, Phone: +82-2-3408-3764, FAX: +82-2-3409-3764, Email: djoh@sejong.ac.kr.
References
- Bagis H., Mercan H.O. Effect of chemically defined culture medium supplemented with β-mercaptoethanol and amino acids on implantation and development of different stage in vivo- or in vitro-derived mouse embryos. Mol. Reprod. Develop. 2004;69:52–59. doi: 10.1002/mrd.20120. [DOI] [PubMed] [Google Scholar]
- Bibila T.A., Robinson D.K. In pursuit of the optical fed-batch process for monoclonal antibody production. Biotechnol. Progr. 1995;11:1–13. doi: 10.1021/bp00031a001. [DOI] [PubMed] [Google Scholar]
- Chen Z., Iding K., Lütkemeyer D., Lehmann J. A low-cost chemically defined protein free medium for a recombinant CHO cell line producing prothrombin. Biotechnol. Lett. 2000;22:837–841. [Google Scholar]
- Ducommun P., Ruffieux P., Stockar U.V., Marison I. The role of vitamins and amino acids on hybridoma growth and monoclonal antibody production. Cytotechnology. 2001;37:65–73. doi: 10.1023/A:1019956013627. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ishaque A., Al-Rubeai M. Role of vitamins in determining apoptosis and extent of suppression by bcl-2 during hybridoma cell culture. Apoptosis. 2002;7:231–239. doi: 10.1023/A:1015343616059. [DOI] [PubMed] [Google Scholar]
- Geshi M., Yonai M., Sakaguchi M., Nagai T. Improvement of in vitro co-culture systems for bovine embryos using a low concentration of carbon dioxide and medium supplemented with 12-mercaptoethanol. Theriogenology. 1999;51:551–558. doi: 10.1016/S0093-691X(99)00009-6. [DOI] [PubMed] [Google Scholar]
- Kim E.J. Development of a Serum-free Medium for the Production of Recombinant Proteins from CHO Cells Using Statistical Design. South Korea: KAIST; 1998. [DOI] [PubMed] [Google Scholar]
- Lao M.S., Toth D. Effects of ammonium and lactate on growth and metabolism of a recombinant Chinese Hamster Ovary cell culture. Biotechnol. Progr. 1997;13:688–691. doi: 10.1021/bp9602360. [DOI] [PubMed] [Google Scholar]
- Lee G.M., Kim E.J., Kim N.S., Yoon S.K., Ahn Y.H., Song J.Y. Development of a serum-free medium for the production of erythropoietin by suspension culture of recombinant Chinese hamster ovary cells using a statistical design. J. Biotechnol. 1999;69:85–93. doi: 10.1016/S0168-1656(99)00004-8. [DOI] [PubMed] [Google Scholar]
- Lee J.C., Chang H.N., Oh D.J. Recombinant antibody production by perfusion cultures of rCHO cells in a depth filter perfusion system. Biotechnol. Progr. 2005;21:134–139. doi: 10.1021/bp0497942. [DOI] [PubMed] [Google Scholar]
- Mercier G., Turque N., Schumacher M. Rapid effects of Triiodothyronine on immediate-early gene expression in Schwann. Cells GLIA. 2001;35:81–89. doi: 10.1002/glia.1073. [DOI] [PubMed] [Google Scholar]
- Morris A.E., Schmid J. Effects of Insulin and LongR3 on serum-Free Chinese Hamster Ovary Cell cultures expressing two recombinant proteins. Biotechnol. Progr. 2000;16:693–697. doi: 10.1021/bp0000914. [DOI] [PubMed] [Google Scholar]
- Oh D.J., Choi S.K., Chang H.N. High-density continuous cultures of hybridoma cells in a depth filter perfusion system. Biotechnol. Bioeng. 1994;44:895–901. doi: 10.1002/bit.260440805. [DOI] [PubMed] [Google Scholar]
- Okamoto T., Tani R., Yabumoto M., Sakamoto A., Takada K., Sato G.H., Sato J.D. Effects of insulin and transferrin on the generation of lymphokine-activated killer cells in serum-free medium. J. Immunol. Methods. 1996;195:7–14. doi: 10.1016/0022-1759(96)00081-6. [DOI] [PubMed] [Google Scholar]
- Pannunzio P., Hazell A.S., Pannunzio M., Rao K.V., Butterworth R.F. Thiamine deficiency results in metabolic acidosis and energy failure in cerebellar granule cells: an in vitro model for the study of cell death mechanisms in Wernicke’s Encephalopathy. J. Neurosci. Res. 2000;62:286–292. doi: 10.1002/1097-4547(20001015)62:2<286::AID-JNR13>3.0.CO;2-0. [DOI] [PubMed] [Google Scholar]
- Ross T.K., Xu F.Y., Taylor W.A., Hatch G.M. Differential effects of chloroquine on cardiolipin biosynthesis in hepatocytes and H9c2 cardiac cells. Mol. Cellular Biochem. 2000;207:115–122. doi: 10.1023/A:1007066903073. [DOI] [PubMed] [Google Scholar]
- Schröder M., Matischak K., Friedl P. Serum- and protein-free media formulations for the Chinese hamster ovary cell line DUKXB11. J. Biotechnol. 2004;108:279–292. doi: 10.1016/j.jbiotec.2003.12.005. [DOI] [PubMed] [Google Scholar]
- Stoll T.S., Miihlethaler K., Stockar U.V., Marison I.W. Systematic improvement of a chemically-defined protein-free medium for hybridoma growth and monoclonal antibody production. J. Biotechnol. 1996;45:111–123. doi: 10.1016/0168-1656(95)00153-0. [DOI] [PubMed] [Google Scholar]
- Sung Y.H., Lim S.W., Chung J.Y., Lee G.M. Yeast hydrolysate as a low-cost additive to serum-free medium for the production of human thrombopoietin in suspension cultures of Chinese hamster ovary cells. Appl. Microbiol Biotechnol. 2004;63:527–536. doi: 10.1007/s00253-003-1389-1. [DOI] [PubMed] [Google Scholar]
- Sunstrom N.S., Gay R.D., Wong D.C., Kitchen N.A., DeBoer L., Gray P.P. Insulin-like growth factor-I and Transferrin mediate growth and survival of Chinese Hamster Ovary cells. Biotechnol. Progr. 2000;16:698–702. doi: 10.1021/bp000102t. [DOI] [PubMed] [Google Scholar]
- Takagi M., Hia H.C., Jang J.H., Yosida T. Effects of high concentrations of energy sources and metabolites on␣suspension culture of Chinese Hamster Ovary Cells producing Tissue Plasminogen Activator. J. Biosci. Bioeng. 2001;91:515–521. doi: 10.1263/jbb.91.515. [DOI] [PubMed] [Google Scholar]
- Trentin A.G., Alvarez-Silva M. Thyroid hormone regulates protein expression in C6 glioma cells Brazilian. J. Med. Biol. Res. 1998;31:1281–1284. doi: 10.1590/s0100-879x1998001000008. [DOI] [PubMed] [Google Scholar]