Multiplication of Swiss 3T3 cells in a serum-free medium

Gently trypsinized Swiss 3T3 cells inoculated into medium MCDB 402 attach readily to polylysine-coated surfaces and remain viable for several days in the absence of exogenously added protein. Short-term multiplication under defined conditions can be obtained by supplementing the MCDB 402 with fibroblast growth factor (FGF), insulin (INS), and dexamethasone (DEX). Addition of bovine plasma fibronectin further improves attachment and viability. This system does not require initial plating in serum or the addition of poorly defined extracts for cellular attachment or for multiplication. In the complete system minus FGF, cells plated at a low density attach to the culture surface and become quiescent. The addition of FGF or PDGF 48–72 h after plating stimulates a high level of DNA synthesis during the following 24 h. EGF also stimulates DNA synthesis in these cells, but to a lesser extent. Insulin and dexamethasone are not needed for the initial DNA synthesis response to FGF, but are needed for continuing multiplication over a period of several days. This system provides a means for studying the effects of specific mitogens on Swiss 3T3 cells in the absence of undefined supplements, and without complications due to density-dependent inhibition.