INFLUENCE OF SERUM COMPONENTS ON GROWTH AND MUTATION OF CHINESE-HAMSTER CELLS IN MEDIUM CONTAINING 8-AZAGUANINE

Low concentrations (.ltoreq. 20 .mu.g/ml) of 8-azaguanine [A) were 1000-fold more toxic to V79 Chinese hamster cells in medium containing 10% dialyzed fetal calf serum [DS] than in medium containing 10% undialyzed serum [US]. Serum enzyme activity that converted AG to nontoxic 8-azaxanthine degraded AG at the same rate, whether or not the serum was dialyzed. Cytotoxicity results similar to those obtained with US were produced in medium containing DS and 2.5 .mu.g of hypoxanthine (HX)/ml (DSH). Serum HX was considered to be responsible for the relatively low cytotoxicity of AG in medium containing US. Colonies that arose in medium containing AG were isolated and characterized. Those that remained resistant to AG (40 .mu.g/ml) and sensitive to aminopterin in the presence of HX and thymidine (HAT) were considered mutants; nonmutants were sensitive to AG and resistant to HAT. Colonies isolated from medium containing DSH or US and low concentrations of AG were not mutants, but those from medium containing high concentrations (.gtoreq. 30 .mu.g/ml) of AG were mutants. Spontaneous and N-methyl-N''-nitro-N-nitrosoguanidine [MNNG] induced mutants were detectable in medium containing DSH without replating the cells prior to adding AG (.gtoreq. 30 .mu.g/ml), but to detect MNNG induced mutations in medium containing DS replating was essential. In DS, the mutation frequency increased as an exponential function of the toxicity of MNNG, but remained 2 orders of magnitude lower than the induced mutation frequencies that occurred in DSH. HX, in DSH or US, produced profound effects, other than interference with AG toxicity, that distort the results of mutagenesis assays. To study mutation using AG resistance as the endpoint, it was essential to use dialyzed serum.