A Qualitative and Quantitative Assay for Cells Lacking Postconfluence Inhibition of Cell Division: Characterization of This Phenotype in Carcinogen-treated Syrian Hamster Embryo Cells in Culture1

We have developed a qualitative and quantitative assay system for detecting cells lacking postconfluence inhibition of cell division (contact insensitivity, CS~) in golden Syrian ham ster embryo cells in culture by measuring the number of cells able to form colonies on a lethally irradiated, confluent monolayer of a contact-sensitive established cell line. A subpopulation in normal low-passage cultures of golden Syrian hamster embryo cells temporarily exhibits this CS~ phenotype at very low frequency (~4 x 10~3) but quickly loses the property within a few passages in vitro. This phenotype is invariably exhibited by various tumorigenic cell lines at very high fre quency (7 to 50 x 10~2) and appears to correlate with the anchorage-independent growth phenotype. The temporal ac quisition of the CS~ phenotype by tertiary-passage golden Syrian hamster embryo cells following exposure to A/-methylA/'-nitro-A/-nitrosoguanidine was examined. Cells with a stably heritable CS~ phenotype are detected after approximately 20 posttreatment population doublings. In contrast, anchorageindependent cells are not detected until 35 to 95 posttreatment population doublings. These CS~ cells appear to be preneoplastic cells, since clonally isolated CS~ cells did not exhibit anchorage-independent growth until after further passaging in vitro. The results suggest that acquisition of the CS~ phenotype represents an early stage in neoplastic progression.