The in vivo alkaline comet assay is used for the detection of DNA stand breaks in cells or nuclei isolated from multiple tissues of animals, usually rodents. The comet assay has been widely used within genetic toxicology for many years and following an extensive validation trial performed over 6 years OECD have published a test guideline. In line with the 3Rs requirements the comet assay can be integrated with other toxicological studies, e.g. repeated dose toxicity studies or the endpoint can be combined with other genotoxicity endpoints, such as the in vivo mammalian erythrocyte micronucleus assay.
The assay has been extensively validated in somatic tissues of rats – stomach and liver. The rationale behind this is that the stomach is usually the first site of contact for substances after oral exposure, and the liver as it is the most active organ in metabolism of substances and is also frequently a target organ for carcinogenicity. The technique in principal can be extended to any tissue from which a single cell/nuclei suspension can be derived, however caution is placed on using this test method for detecting DNA effects in germ cells
Verification of laboratory proficiency in conducting the comet assay must be demonstrated, both in terms of experimental competency and also in the collection of negative control data to demonstrate reproducibility of negative data responses and positive control data to show the ability to detect such responses. Historical control ranges and distributions must be established for relevant tissues and species that are being investigated.
The comet test guidelines can be viewed on the OECD website (http://www.oecd-ilibrary.org/environment/oecd-guidelines-for-the-testing-of-chemicals-section-4-health-effects_20745788).
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