Conditionally adherent growth of serum-independent CHO cells for automated drug screening and biopharmaceutical production.
SSF3 is a CHO cell line adapted for growth in protein-free medium. It grows in suspension unless serum-derived attachment factors such as vitronectin are added to the medium. Serum-independent cell lines, which adhere to the substrate after induction with dexamethasone or constitutively, were created by transfection with a human vitronectin gene under control of the mouse mammary tumor-virus promoter. Substrate attachment and SSF3VN-cell spreading could be prevented with an RGD peptide (arginine-glycine-aspartic acid) confirming that attachment is mediated by an intregrin receptor. Hormone-inducible attachment could be blocked by glucocorticoid antagonist promegestone. All steps in the isolation of stable transfected SSF3VN cell lines could be done in a chemically defined medium avoiding the risk of introduction of serum-derived infectious agents. SSF3VN cells could be grown in protein-free medium in solid-phase large-scale bioreactors. Application in microplates as used in high-throughput screening was demonstrated in an assay of Ca(2+) release from internal stores induced by agonist-binding to recombinant human metabotropic glutamate receptor hmGluR1b.
Gandor
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EugeneX Biotechnologies GmbH, Lauchefeld 31, CH-9548 Matzingen, Switzerland. m.o.zang-gandor@eugenex.com
Zang-Gandor
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Marangi
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D000818:Animals; D001702:Biopharmaceutics / Q000379:methods; D019149:Bioreactors; D016466:CHO Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D002448:Cell Adhesion; D002455:Cell Division; D006224:Cricetinae; D016895:Culture Media, Serum-Free; D004353:Drug Evaluation, Preclinical / Q000379:methods; D006801:Humans; D051379:Mice; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection; D019096:Vitronectin / Q000096:biosynthesis / Q000235:genetics
D016895:Culture Media, Serum-Free; D011994:Recombinant Proteins; D019096:Vitronectin