Abstract
Transformation of rat hepatocytes by simian virus 40 in chemically defined medium was examined. When hepatocytes plated on collagen-coated plates were infected with simian virus 40, colonies of replicating cells appeared as early as 40 days after infection, whereas no colonies appeared in control cultures. Medium from 85% of the transformed cultures contained albumin. When collagen was eliminated and hepatocytes were plated on Primaria plastic cell culture dishes, transformation occurred; medium from 86% of the transformed cultures contained albumin but the maximum albumin level secreted per culture was only 62% of that produced by cultures on collagen-coated plates. Quantitative assays for transformation were established. Transformation was linear after infection with 2-50 plaque-forming units of virus per hepatocyte, and the transformation frequency was the same on the two plating surfaces. An immuno-overlay technique made it possible to identify, purify, and determine the morphology of the albumin-producing cells. When ornithine was substituted for arginine in the medium, the transformation frequency decreased markedly while the percentage of colonies producing albumin increased from 30% to 100%. We conclude that we have defined an assay for quantifying transformation of a normal hepatocyte population and for identifying and enumerating epithelial liver cell transformants that produce albumin.