The use of serum-free medium for the production of functionally active humanised monoclonal antibody from NS0 mouse myeloma cells engineered using glutamine synthetase as a selectable marker

• Published in: Cytotechnology (Dordrecht) 1995-01, Vol.18 (3), p.207-217
• Main Authors: KEEN, M. J, HALE, C
• Format: Article
• Language: English
• Subjects: Animal cells; Biological and medical sciences; Biotechnology; Establishment of new cell lines, improvement of cultural methods, mass cultures; Eukaryotic cell cultures; Fundamental and applied biological sciences. Psychology; Methods. Procedures. Technologies
• ISSN: 0920-9069; 1573-0778
• DOI: 10.1007/BF00767768

A protein-free growth medium (W38 medium) had previously been developed for the NS0 mouse myeloma cell line which is cholesterol-auxotrophic. This paper describes the development of a protein-free growth medium for NS0 cells expressing humanised monoclonal antibody using GS (glutamine synthetase) as a selectable marker. Several GS-engineered NS0 cell lines expressing humanised monoclonal antibody grew in a modification of W38 medium which maintained GS-selection, supplemented with cholesterol, phosphatidylcholine and β-cyclodextrin. Further studies showed that additional glutamic acid, asparagine, ribonucleosides and choline chloride improved cell growth. Amino acid analysis identified a number of amino acids that were being depleted from the culture medium. NS0 cell lines 9D4 and 2H5 expressing CAMPATH-1H(*) were adapted to enable them to grow serum-free in the absence of cholesterol and β-cyclodextrin. Cholesterol-independent 9D4 (9D4.CF) cells grown in shake flask culture using an enriched protein-free medium (WNSD medium), supplemented with human recombinant insulin (Nucellin), reached a maximum cell density to 1.86×10(6) cells ml(-1) producing 76.6 mg l(-1) of antibody. CAMPATH-1H antibody produced using serum-free medium was found to be functionally activein vitro in the Antibody Dependant Cellular Cytotoxicity (ADCC) assay.