TM2020-01 Report on the Full Test Submission Assessment
The RS Comet assay is an in vitro genotoxicity assay, presented as an adaptation of the known alkaline comet assay, which measures DNA strand breaks in eukaryotic cells, and for which an OECD in vivo mammalian Test Guideline is currently available (OECD TG 489). The RS Comet assay has been designed in a three-dimensional reconstructed human skin (RHS) model. The method uses specifically the Phenion® Full-Thickness Skin Model which is composed of human primary keratinocytes and fibroblasts cultured under air-liquid-interface (ALI) conditions on a collagen scaffold. The Phenion® Full-Thickness Skin Model resembles the fully differentiated epidermis and the underlying dermis.
The RS Comet assay has been presented as a relevant in vitro test method to investigate DNA damage after dermal exposure and, particularly, for substances under regulations that prohibit or limit in vivo testing, to follow up positive results from the standard in vitro genotoxicity test battery. The test method has undergone a full validation process thus, the Test Submitter has requested EURL ECVAM to consider the RS Comet assay for peer-review.
The full Test Submission, received on May 2021, was completed in all its parts. EURL ECVAM acknowledged that the submission included a detailed test protocol (SOP) together with data annexes and bibliography describing the validation study, the properties of full-thickness skin model, and strategic use of the RS Comet assay, including in regulatory context i.e., under Cosmetics Regulation (described in paragraph 3.9. and 4 of the Test Submission Template (TST)).
EURL ECVAM was able to assess the test method’s readiness to enter peer review by evaluating the completeness and quality of the information provided for each of the following relevant modules: test definition, within-laboratory reproducibility (WLR), transferability, between-laboratory reproducibility (BLR), predictive capacity and applicability domain. Suggestions for performance standards (PS) were not provided at this stage. Nevertheless the lack of information did not affect the EURL ECVAM assessment.
The Test Submitter has highlighted that the RS Comet assay can be used together with the RSMN (reconstructed skin micronuclei assay) in order to cover all genotoxicity endpoints that usually need to be addressed for regulatory purposes (gene mutation, clastogenicity, and aneugenicity).
Based on the mechanistic and biological relevance of the test method and provided information, EURL ECVAM concludes that the RS Comet assay is ready to undergo to peer review.
MENNECOZZI Milena;
CORVI Raffaella;
BARROSO João;
2025-12-23
Publications Office of the European Union
JRC141979
978-92-68-27156-8 (online),
OP KJ-01-25-268-EN-N (online),
https://publications.jrc.ec.europa.eu/repository/handle/JRC141979,
10.2760/2040379 (online),
