Bacterial Reverse Mutation Test (Ames)

According to OECD Guidelines for Testing of Chemicals, Test No. 471, the bacterial reverse mutation test (Ames) is performed with histidine-dependent Salmonella typhimurium strains, or Escherichia coli to detect permanent gene mutations, such as substitutions, additions or deletions of one or several DNA base pairs caused by new chemicals and drug candidates.

STEMart provides bacterial reverse mutation test to assess the potential mutagenic activity of leachable from a medical device. This assay is available under either GLP (Good Laboratory Practice) or Non-GLP conditions.

Standard

OECD 471

Test Strains

• S. typhimurium strains carrying point mutations in histidine operon: TA97, TA97a, TA98, TA100, TA102, TA1535, TA1537 and TA1538;
• E. coli strains carrying point mutations in tryptophan operon: E. coli uvrA, E. coli pKM101 and E. coli uvrA (pKM101).

At least five bacteria strains shall be used in Ames test. The number and set of strains can be determined based on the regulatory requirements.

Test Method

Cell suspensions of five bacterial strains are exposed to the test substance (liquid or solid) in the presence and in the absence of an appropriate exogenous metabolic activation system (chemically induced rat liver S9 fraction).

Using plate incorporation technology or preincubation method, the bacteria are cultured in histidine- or tryptophan-deficient media for two or three days at 37°C.

Negative vehicle control and positive controls are included in each study.

Revertant colonies are counted and are compared to that of the control plates.

Fig.1 Ames test procedure

Result Interpretation

Test sample with mutagenic potential makes auxotrophic strain regain the ability to synthesize histidine/tryptophan. The mutagenicity of test sample is proportional to number of colonies observed.

• Positive result: A more than two-fold increase of colonies over the mean negative control value;
• Negative result: No dose-related increase in all five tester strains.

Final Report

The final report including information on the methodology, raw data, analysis, and interpretation of the results will be provided for customer.

Advantages

• Robust, rapid, low cost and easy to perform;
• Suitable mutants can be detected in a wide range of bacteria with high sensitivity;
• Gold standard assay for prediction of human carcinogenic potential of a chemical by health regulatory authorities;
• First screen assay for determination of the mutagenic potential of new chemicals and drugs;
• The specificity of the test strains can offer some valuable information about the types of mutations induced by genotoxic agents;
• Detection and distinction between direct acting mutagens and pro-mutagens that need metabolization.

If you have additional questions about bacterial reverse mutation test (Ames), or would like to find out more about our services, please feel free to contact us.

References

• Mortelmans, K., et al. "The Ames Salmonella." Microsome Mutagenicity (2000).
• Maron, Dorothy M., et al. "Revised methods for the Salmonella mutagenicity test." Mutation Research/Environmental Mutagenesis and Related Subjects 113.3-4: 173-215 (1983).
• OECD, Test No. "471: bacterial reverse mutation test." OECD Guidelines for the Testing of Chemicals, Section 4 (1997).