Improved medium and culture conditions for clonal growth with minimal serum protein and for enhanced serum-free survival of Swiss 3T3 cells.

Improved culture conditions have been developed that will support clonal growth of Swiss mouse embryo 3T3 cells at concentrations of serum protein at low as 125 micrograms/ml. Survival of the cells under completely protein-free conditions also is enhanced greatly. The improvements that made these results possible include: (a) use of medium MCDB 402, which was developed specifically for Swiss 3T3 cells by adjusting the concentrations of all components of Dulbecco's modified Eagle's medium to optimum values for clonal growth with minimal serum protein and by adding other nutrients such as trace elements and "nonessential" amino acids that were not in the original formula; (b) use of culture surfaces that are coated with a positively charged polymer, poly-D-lysine; and (c) use of gentle low temperature trypsinization technique that minimizes cellular damage and the need to neutralize residual trypsin.