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Kuwana Y, Funayama K, Miyaji H, et al. Production of recombinant granulocyte colony-stimulating factor by knocking into the active immunoglobulin heavy chain gene locus in the hybridoma cell line. Cytotechnology. 2001;37(3):133-41doi: 10.1023/A:1020585320775.
Kuwana, Y., Funayama, K., Miyaji, H., Hasegawa, M., Yoshida, H., & Itoh, S. (2001). Production of recombinant granulocyte colony-stimulating factor by knocking into the active immunoglobulin heavy chain gene locus in the hybridoma cell line. Cytotechnology, 37(3), 133-41. https://doi.org/10.1023/A:1020585320775
Kuwana, Y, et al. "Production of recombinant granulocyte colony-stimulating factor by knocking into the active immunoglobulin heavy chain gene locus in the hybridoma cell line." Cytotechnology vol. 37,3 (2001): 133-41. doi: https://doi.org/10.1023/A:1020585320775
Kuwana Y, Funayama K, Miyaji H, Hasegawa M, Yoshida H, Itoh S. Production of recombinant granulocyte colony-stimulating factor by knocking into the active immunoglobulin heavy chain gene locus in the hybridoma cell line. Cytotechnology. 2001 Nov;37(3):133-41. doi: 10.1023/A:1020585320775. PMID: 19002916; PMCID: PMC3449790.
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Cytotechnology. 2001 Nov;37(3):133-41. doi: 10.1023/A:1020585320775.

Production of recombinant granulocyte colony-stimulating factor by knocking into the active immunoglobulin heavy chain gene locus in the hybridoma cell line.

Cytotechnology

Y Kuwana, K Funayama, H Miyaji, M Hasegawa, H Yoshida, S Itoh

Affiliations

  1. Tokyo Research Laboratories, Kyowa Hakko Kogyo, Co. Ltd., 3-6-6 Asahi-machi, 194-8533, Tokyo, Japan, [email protected].

PMID: 19002916 PMCID: PMC3449790 DOI: 10.1023/A:1020585320775

Abstract

The hybridoma cell line KM50 originally produces a monoclonal antibody at a concentration of approximately 40 mg ml(-1) in ascites. To investigate the possibility to apply this expression system to the production of useful proteins, the cDNA encoding human granulocyte colony-stimulating factor was inserted by homologous recombination into just downstream of the promoter of the active immunoglobulin heavy chain gene of KM50. Site directed integration of targeting DNAs resulted in the disruption of expression of the immunoglobulin heavy chain proteins with a frequency of 1 in 10 approximately 100 G418-resistance transfectants. One of the monoclonal antibody-deficient transfectants produced25 ng ml(-1) of granulocyte colony-stimulating factor in the supernatant of its cell culture the number of molecules of which corresponds to that of the monoclonal antibody originally produced by KM50. However, when this transfectant was injected intraperitoneally, it produced only a 9 mug ml(-1) concentration of granulocyte colony-stimulating factor in ascites, which is approximately 3 orders of magnitude less than the monoclonal antibody. This method may be applicable to production of other recombinant proteins, although further optimization in the conditions of production would be needed in order to reach much higher yields.

References

  1. Proc Natl Acad Sci U S A. 1989 Nov;86(21):8507-11 - PubMed
  2. Cytotechnology. 1991 Mar;5(3):223-31 - PubMed
  3. J Biochem. 1987 May;101(5):1307-10 - PubMed
  4. Gene. 1999 Feb 4;227(1):21-31 - PubMed
  5. Immunology. 1995 Jan;84(1):159-63 - PubMed
  6. Cell. 1986 Jan 31;44(2):283-92 - PubMed
  7. Mol Biother. 1992 Jun;4(2):70-6 - PubMed
  8. J Immunol. 1994 Jan 15;152(2):695-704 - PubMed
  9. Annu Rev Immunol. 1991;9:373-98 - PubMed
  10. Gene. 1998 Nov 19;222(2):279-85 - PubMed
  11. J Immunol Methods. 1999 May 27;225(1-2):185-9 - PubMed
  12. Proc Natl Acad Sci U S A. 1980 Apr;77(4):2138-42 - PubMed
  13. Gene. 1984 Feb;27(2):223-32 - PubMed
  14. J Biol Chem. 1991 Oct 25;266(30):19867-70 - PubMed
  15. Jpn J Cancer Res. 1987 Nov;78(11):1179-81 - PubMed
  16. Eur J Immunol. 1995 Mar;25(3):792-7 - PubMed
  17. Annu Rev Immunol. 1991;9:159-91 - PubMed
  18. Mol Cell Biol. 1997 May;17(5):2658-68 - PubMed
  19. Proc Natl Acad Sci U S A. 1986 Aug;83(16):6075-9 - PubMed
  20. J Clin Oncol. 1998 May;16(5):1861-8 - PubMed
  21. Biochem Biophys Res Commun. 1989 Feb 28;159(1):103-11 - PubMed
  22. Genes Dev. 1988 Nov;2(11):1353-63 - PubMed
  23. Clin Exp Pharmacol Physiol. 1998 Jul-Aug;25(7-8):568-71 - PubMed
  24. Nature. 1986 Jan 30-Feb 5;319(6052):415-8 - PubMed
  25. Gene. 1997 Apr 1;188(2):191-8 - PubMed
  26. Mol Gen Genet. 1997 Nov;256(5):499-508 - PubMed

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