Effects of incubation in an atmosphere of 20% CO2 in air on the Syrian hamster embryo clonal transformation assay.

An atmosphere containing 10% CO2 has been generally accepted as optimal for the growth of Syrian hamster embryo cells in a clonal transformation assay. Data presented in this paper show that 10% CO2 may not be the optimum environment for this assay. Using 10 or 20% (analytically measured) CO2 in air (1 atm pressure), hamster embryo cell pools were examined for clonal growth characteristics and transformability using five known carcinogens and a single noncarcinogenic compound. At 10% CO2, only 2 of 11 pools were transformed by the five carcinogens but not by the noncarcinogen. At 20% CO2, six of seven pools were transformed by the five carcinogens and not by the noncarcinogen. Further, the transformation frequencies were found to be greater in cultures incubated in an atmosphere consisting of 20% CO2 in air. The data also show that 20% CO2 increased the cloning efficiency of these cells. A comparison of the 10 and 20% CO2 data to results reported from other laboratories suggests that conflicting interlaboratory results with this assay system may be due, in part, to variations of CO2 concentrations. In some instances, the CO2 levels indicated by incubator flow meters vary considerably from analytically determined CO2 values. To prevent these CO2 discrepancies and their resultant effects on transformation and cloning efficiency, methods for monitoring the CO2 environment other than flow meters are recommended. The observation of increased cloning efficiencies and transformation rates strongly suggests that culture incubation at 20% CO2 is a preferred environment for the conduct of this assay.