Growth of NS0 cells in protein-free, chemically defined medium

Many hybridoma and recombinant myeloma cell lines have been successfully ad apted to growth in protein-free media. Compared with serum-supplemented med ia, use of protein-fi ee media promotes superior cell growth and protein ex pression and facilitates downstream purification of the expressed product. Owing to its sterol auxotrophy, the NS0 myeloma is normally grown in either a serum-supplemented medium or a serum-free medium supplemented with an an imal-derived lipoprotein. CD Hybridoma Medium (a protein-free, chemically d efined formulation) grows many cell lines that do not exhibit lipid depende nce, but this medium does not support growth of NS0 cells without further l ipid supplementation. We tested several commercially available lipid supple ments in CD Hybridoma Medium, including bovine EX-CYTE VLE. None of the tes ted supplements supported longterm growth of NS0 cells in CD Hybridoma Medi um. Sustained long-term growth of NS0 cells was achieved in CD Hybridoma Me dium supplemented with various animal- or plant-derived lipids complexed wi th cyclodextrin. NS0 cells adapted to CD Hybridoma Medium supplemented with cyclodextrin-lipid complex reached peak cell densities that were more than double those observed in serum-supplemented medium and were cultured for m ore than 15 passages. These cultures were also successfully cryopreserved a nd recovered in this defined medium. Through the use of cyclodextrin-based additives to CD Hybridoma Medium, it is possible to solubilize significant quantities of sterols and other lipids and to maintain a protein-free, chem ically defined cultivation environment for NS0 cells. The culture system ca n be kept entirely free of animal-derived components if the supplement is m ade with plant-derived or synthetic Lipids.