Factors affecting the performance of the styles cell transformation test

The effects of certain factors on the performance of the cell transformation test of Styles were examined by testing the demonstrability of transformation of cells of a subclone of BHK21 C13 in response to treatment with 4-nitroquinoline-1-oxide, N-methyl-N'-nitro-N-nitrosoguanidine, benzo[a]pyrene and 2-acetamidofluorene. An important requirement for success was supplementation of the soft agar medium with a serum which supported microcolony formation by a high proportion of the cells. Increasing the concentration of an unsuitable serum improved the results obtained; this suggests that the serum was inadequate rather than inhibitory. Alteration of the concentration of S-9 fraction used to activate the precarcinogens benzo[a]pyrene and 2-acetamidofluorene had little effect on the induction of transformation by either. A clearer distinction between the transformation frequencies for control and treated cells was usually obtained when 500 microm rather than 200 microm was the minimum diameter set for definition of transformation. It is suggested that, to assess the validity of results obtained with compounds which appear to induce transformation only at highly toxic levels, transformation frequencies for treated cells should be compared with those for control cells seeded at comparable densities.