An Introduction To The Ames Test

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The Ames test is a method commonly used by specialist CRO companies like Gentronix, to test if a particular chemical can cause mutations within the DNA of the test organism through the use of bacteria. Used to determine the mutagenic potential of chemical compounds, the Ames Test is a commonly used regulatory toxicology assay. If the test gives a positive result, it may indicate that the chemical is mutagenic and thus could act as a carcinogen, since cancer is usually linked to mutation. The test can also serve as a quick and easy assay to determine the genotoxic carcinogenic potential of a compound – a standard carcinogen assay on rats and mice is typically very time-consuming (this can take two to three years to complete) and not cost-efficient. It will not detect non-genotoxic carcinogens though. However, it is also noted that false negatives and false positives can be seen through this.

The test works by using a Salmonella Typhimurium histidine (His) reversion system, involving a mutation of the histidine locus in the genome of multiple strains. This test is conducted in compliance with the Organization for Economic Cooperation and Development (OECD) 471 Guideline for Testing of Chemicals: Bacterial Reverse Mutation Test. Abbreviated assays can be used for some specific situations, for example, the modified Ames Oil test and Mutagenic Activity of Water Extract test (AS/NZS 4020).

Based on the principle of back mutation or reverse mutation, the test can also be referred to as a bacterial reverse mutation assay.

  • Test organism: Ames tests use multiple strains of bacteria (Salmonella, E. coli) that carry specific mutations. Because these mutations are not all carried in the same place in the genes that encode histidine, multiple mutagenic mechanisms can be tested for.
  • Culturing His- Salmonella in media containing test chemicals can result in mutation in the histidine encoding gene – this means that they can regain the ability to synthesize histidine (His+), allowing colonies to grow in the absence of histidine. Such chemicals are responsible for reverting the mutation are known as a mutagen. Therefore, this Ames test can be used to test for the mutagenic ability of a vast array of chemicals.

The Ames Salmonella mutagenicity assay is the most common short-term in vitro genotoxicity test. The assay employs specific strains of the bacterium Salmonella typhimurium which have been mutated at genetic loci precluding the biosynthesis of the amino acid histidine which is needed for reproduction and growth.

The mutagenicity assay depends upon treating the bacteria with test material across an array of doses immediately below the concentration showing vital toxicity to bacteria. The treated bacteria are thus grown on agar plate deficient in histidine.

The original mutated Salmonella (histidine-dependent) is not able to form visible colonies under these growing conditions unless they were to undergo a second mutation and change back to histidine-independent. Once the test material is treated, the quantity of such revertant colonies per agar plate can be used to indicate the mutagenic potency of the test material.

Specific Applications of the Ames Test

The modified Ames test can be used to study the potential of short-term in vitro genotoxicity by using a particular type of Salmonella Typhimurium which has been genetically engineered to preclude the ability for amino acid histidine synthesis. It can also be used as a screening technique which detects the presence of potential dermal carcinogens in virgin base oils which are used in the creation of metalworking fluids. Another common use of the Ames test is to test waters from plumbing products in contact with drinking water.