Abstract:

OBJECTIVE: We developed a standard Ames fluctuation test in our laboratory and lay the foundation for its application in detecting the genetic toxicity of compounds. METHODS：We used four positive compounds in the test，Dexon，SA，CP and 2-AF. We compared two ways of enrichment culture and two indicators to eatablish the sandand percedure. We adopted 0.1，1，10 μg/mL Dexon and 0.01，0.1，1 μg/mL SA in the non-activated condition with their menstruum as negative control；10，100，1 000 μg/mL CP and 0.1，1，10 μg/mL 2-AF in the activated condition with their menstruum as negative control. We recorded the number of positive wells，adjusting the dose of the positive compounds accordingly，until there was a significant difference (P<0.01) between the positive group and the solvent compared group. After determining the dose of the positive control compounds，we repeated the test at the same dose 3 times，to ensure that the test system was stable and reliable. RESULTS：We determined the doses of the four positive compounds for the Ames fluctuation test in our laboratory，Dexon 3 μg/mL，SA 0.05 μg/mL，CP 100 μg/mL and 2-AF 30 μg/mL. In the non-activated condition 3 μg/mL Dexon and 0.05 μg/mL SA of the 3 96-well plates were significantly different from the solvent control group，exhibiting strong mutagenic effect to the TA100(P<0.01). Similarly，100 μg/mL CP and 30 μg/mL 2-AF in the activated condition of the 3 96-well plates were significantly different from the solvent control group，also showing strong mutagenic effect to the TA100(P<0.01). We also improved the test method during the experiment，including sandardizing the doses of test substance，improving enrichment culture method and determining the indicator. CONCLUSION：We established the Ames fluctuation test in our laboratory，confirmed that the known positive compounds were good and stable，and we optimized the experimental conditions in the study.

Key words: Ames fluctuation test, mutagenicity test, standardization, genetic toxicity