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Cytotechnology

, Volume 17, Issue 3, pp 153–163 | Cite as

Development of a serum-free culture medium for the large scale production of recombinant protein from a Chinese hamster ovary cell line

  • Michael J. Keen
  • Nicholas T. Rapson
Article

Abstract

A serum-free medium, WCM5, has been developed for the large scale propagation of CHO (Chinese hamster ovary) cells which express recombinant protein using dihydrofolate reductase as a selectable marker. WCM5 was prepared by supplementing Iscoves medium without lecithin, albumin or transferrin with a number of components which were shown to benefit growth. WCM5 medium contained 5 mg l−1 human recombinant insulin (Nucellin) but was otherwise protein-free. CHO 3D11* cells which had been engineered to express a humanised antibody, CAMPATH*-1H, were routinely grown using serum-containing medium. From a seeding density of 105 cells ml−1, cells grown in static culture with serum reached a maximal cell density of 6.5×105 cells ml−1 after 6 days in culture and produced a maximal antibody concentration of 69 mg l−1 after 11 days in culture. CHO 3D11* cells grown with serum were washed in serum-free medium then cultured in WCM5 medium. Following a period of adaptation the cell growth and product yield was superior to that achieved with serum-containing medium. CHO cells producing CAMPATH-1H grown in an 8000 l stirred bioreactor seeded with 2×105 cells ml−1 reached a maximal viable cell density of 2.16×106 cells ml−1 after 108 h in culture and a maximal antibody concentration of 131.1 mg l−1 after 122 h in culture.

Key words

Insulin ferric citrate Chinese hamster ovary dhfr serum-free 

Abbreviations

CHO

Chinese hamster ovary

dhfr

dihydrofolate reductase

dhfr

dihydrofolate reductase deficient

MTX

methotrexate

H

hypoxanthine

T

thymidine

T/V

trypsin versene

F12

Hams F12 medium

NEAA

non essential amino acids

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Copyright information

© Kluwer Academic Publishers 1995

Authors and Affiliations

  • Michael J. Keen
    • 1
  • Nicholas T. Rapson
    • 1
  1. 1.Biology Research DivisionWellcome Research LaboratoriesBeckenhamUK

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