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Critical issues with the in vivo comet assay: a report of the comet assay working group in the 6th International Workshop on Genotoxicity Testing (IWGT)

Plappert Helbig, Ulla, Speit, Günter, Kojima, Hajime , Burlinson, Brian, Collins, Andrew, Kasper, Peter, Uno, Yoshifumi , Vasquez, Marie, Beevers, Carol, De Boeck, Marlies, Escobar, Patricia, Pfuhler, Stefan and Dan D. Levy, Dan D. (2015) Critical issues with the in vivo comet assay: a report of the comet assay working group in the 6th International Workshop on Genotoxicity Testing (IWGT). Mutation Research , 783 . pp. 6-12.


As a part of the 6th IWGT, an expert working group on the comet assay evaluated
critical topics related to the use of the in vivo comet assay in regulatory genotoxicity testing.
The areas covered were: identification of the domain of applicability and regulatory
acceptance, identification of critical parameters of the protocol and attempts to standardize the
assay, experience with combination and integration with other in vivo studies, demonstration
of laboratory proficiency, sensitivity and power of the protocol used, use of different tissues,
freezing of samples, and choice of appropriate measures of cytotoxicity. The standard
protocol detects various types of DNA lesions but it does not detect all types of DNA damage.
Modifications of the standard protocol may be used to detect additional types of specific DNA
damage (e.g., cross-links, bulky adducts, oxidized bases). In addition, the working group
identified critical parameters that should be carefully controlled and described in detail in
every published study protocol.
in vivo comet assay results are more reliable if they were obtained in laboratories that
have demonstrated proficiency. This includes demonstration of adequate response to vehicle
controls and an adequate response to a positive control for each tissue being examined. There
was a general agreement that freezing of samples is an option but more data are needed in
order to establish generally accepted protocols. With regard to tissue toxicity, the working
group concluded that cytotoxicity could be a confounder of comet results. It is recommended
to look at multiple parameters such as histopathological observations, organ-specific clinical
chemistry as well as indicators of tissue inflammation to decide whether compound-specific
toxicity might influence the result. The expert working group concluded that the alkaline in
vivo comet assay is a mature test for the evaluation of genotoxicity and can be recommended
to regulatory agencies for use.

Item Type: Article
Keywords: in vivo comet assay genotoxicity testing genotoxicity test strategies guideline for the comet assay
Date Deposited: 13 Oct 2015 13:12
Last Modified: 13 Oct 2015 13:12


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