Adaptation of cholesterol-requiring NS0 mouse myeloma cells to high density growth in a fully defined protein-free and cholesterol-free culture medium.

摘要: NS0 has been used as a fusion partner for the production of hybridomas and more recently engineered to produce recombinant protein. A protein-free culture medium, designated W38 previously developed which supported high density growth rat myeloma hybridoma cell lines. cells failed grow in medium number media support other are derived from NS-1 line, is known require exogencus cholesterol. It was found that grew supplemented with phosphatidylcholine, cholesterol, albumin were auxotrophic Protein-free achieved by using β-cyclodextrin replace lipid carrier. The maximal reached this excess 1.5×106 ml−1. supplements precipitated after few days storage at +4°C. In order overcome problem protocol allowed be adapted cholesterol-independent medium. NS0.CF (cholesterol-independent cells) cultured continuously several months. shake flask 2.4×106 ml−1 compared 1.41×106 RPMI 1640 containing 10% foetal bovine serum. readily 1 litre stirred bioreactor Pluronic F68 reaching 3.24×106 cloned limiting dilution giving colonies wells seeded an average 0.32 per 200 μl. This study demonstrated first time cholesterol-requiring mouse line completely defined its subsequent adaptation cholesterol-independence.