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Six mutagenic and/or carcinogenic agents were tested for their ability to cause somatic eye mutation (red sectors on a yellow eye) in Drosophila melanogaster. Results were analysed by the chi-squared method and by the Kolmogorov-Smirnov two-sample (non-parametric) test. Methyl methanesulphonate, cyclophosphamide, acriflavine and procarbazine were active, whereas acetamide and tetracycline failed to elicit a significant response. Thus, relative to other mutagenicity assays, this somatic system seemed to possess no special ability to correctly identify carcinogens because both acriflavine and acetamide were misidentified on a carcinogenicity basis. However, it could be a suitable alternative to the recessive lethal test for mutation, because of its technical simplicity and rapidity, subject to an increase in the size of the data base.

The optimum concentrations of Aroclor-induced rat liver S9 microsomal fraction for the mutagenic activity of the four standard mutagens 2-aminofluorene (2-AF), acriflavine (ACR), benzo[a]pyrene (BP) and cyclophosphamide (CP) were determined in four mutation assays. The four assays were the Ames test using Salmonella typhimurium strain TA100, cycloheximide resistance in the yeast Saccharomyces cerevisiae, the mouse lymphoma TK assay and the human peripheral lymphocyte cytogenetic assay. BP was the only mutagen to be most active at comparable S9 concentrations, of approximately 1%, for all four assays. The optimum S9 concentrations for each of the remaining three mutagens varied substantially between the four assays. ACR was a potent direct-acting mutagen in both mammalian cell assays. The mouse lymphoma TK assay results showed similar optimal values of 1.5% S9 or below for each of the four test agents. The assay with the largest variation of optimal S9 values for the four mutagens was the Ames test in strain TA100, although it also had the widest peaks of activity over the range of S9 concentrations tested. It is likely that the diversity of findings is due to a variety of metabolites affecting the different genetic endpoints that are measured in these assays. Thus from these results it is not possible for bacterial optimization data to be related to other routine in vitro systems. The use of more than one concentration of S9 would contribute useful information.

The definition of a negative result is a problem in genetic toxicology. Here we suggest that a result may be considered biologically unimportant (negative) if it falls within the limits of variation usually found in the negative controls of the particular test. To determine 'usual' variation, we have set 95% confidence limits on three indices of variation, calculated from historical values for duplicate negative control data from several genotoxicity tests. These tests showed four characteristic types of response and the appropriate index of variability was determined for each. Where there was little test-to-test variation in true mean (micronucleus test and metaphase analysis), confidence limits set on the overall distribution of negative controls were the best index of variability. In other assays (Ames, yeast and mouse lymphoma), there was considerable test-to-test variation so that differences between, or ratios of, the members of control duplicates were the preferred measure of variability. This approach can define what is biologically unimportant in terms of the test system. However, no inference can be drawn as to potential importance. Thus the main use is the removal of the positive 'label' from statistically significant results which fall within the usual range of spontaneous variation for the assay under consideration.