The growth requirements of SV40 virus transformed Balb/C‐3T3 cells in serum‐free monolayer culture

The growth requirements of SV40 transformed Balb/c-3T3 cells have been studied in the absence of serum. For growth in serum-free medium, the cells require (i) insulin, (ii) transferrin, and (iii) cis-unsaturated fatty acids added in combination with fatty acid free bovine serum albumin. The growth rate, saturation density, and morphology of cells grown in this serum-free medium are the same as those of cells grown in serum supplemented medium. This mixture also supports the growth of SV40 transformed Swiss-3T3 cells and SV40 transformed primary mouse embryo cells, but does not support the growth of untransformed Balb/c-3T3 cells. The addition of fibronectin to this mixture allows routine subculture, repeated passage, and indefinite propagation of SV40 transformed Balb/c-3T3 cells. Cells grown in this medium for a period of two months retain their ability to induce tumors when injected into athymic nude mice.