1. Inter-laboratory variation in DNA damage using a standard comet assay protocol
- Author
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Lykke, Forchhammer, Clara, Ersson, Steffen, Loft, Lennart, Möller, Roger W L, Godschalk, Frederik J, van Schooten, George D D, Jones, Jennifer A, Higgins, Marcus, Cooke, Vilas, Mistry, Mahsa, Karbaschi, Andrew R, Collins, Amaya, Azqueta, David H, Phillips, Osman, Sozeri, Michael N, Routledge, Kirsty, Nelson-Smith, Patrizia, Riso, Marisa, Porrini, Giuseppe, Matullo, Alessandra, Allione, Maciej, Stępnik, Maciej, Steepnik, Magdalena, Komorowska, João Paulo, Teixeira, Solange, Costa, Laura-Ana, Corcuera, Adela, López de Cerain, Blanca, Laffon, Vanessa, Valdiglesias, Peter, Møller, Farmacologie en Toxicologie, and RS: NUTRIM - R4 - Gene-environment interaction
- Subjects
DNA damage ,Endpoint Determination ,Health, Toxicology and Mutagenesis ,Toxicology ,DNA-formamidopyrimidine glycosylase ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Validation ,Genetics ,Humans ,Inter-laboratory ,Genetics (clinical) ,030304 developmental biology ,Protocol (science) ,Inter-laboratory Variation ,0303 health sciences ,Chromatography ,Ar e Saúde Ocupacional ,Chemistry ,Formamidopyrimidine DNA glycosylase ,3. Good health ,Comet assay ,DNA-Formamidopyrimidine Glycosylase ,030220 oncology & carcinogenesis ,Calibration ,Standard protocol ,Leukocytes, Mononuclear ,Linear Models ,Comet Assay ,Genotoxicidade Ambiental ,Laboratories ,DNA ,Biomarkers ,DNA Damage ,Human - Abstract
There are substantial inter-laboratory variations in the levels of DNA damage measured by the comet assay. The aim of this study was to investigate whether adherence to a standard comet assay protocol would reduce inter-laboratory variation in reported values of DNA damage. Fourteen laboratories determined the baseline level of DNA strand breaks (SBs)/alkaline labile sites and formamidopyrimidine DNA glycosylase (FPG)-sensitive sites in coded samples of mononuclear blood cells (MNBCs) from healthy volunteers. There were technical problems in seven laboratories in adopting the standard protocol, which were not related to the level of experience. Therefore, the inter-laboratory variation in DNA damage was only analysed using the results from laboratories that had obtained complete data with the standard comet assay protocol. This analysis showed that the differences between reported levels of DNA SBs/alkaline labile sites in MNBCs were not reduced by applying the standard assay protocol as compared with the laboratory's own protocol. There was large inter-laboratory variation in FPG-sensitive sites by the laboratory-specific protocol and the variation was reduced when the samples were analysed by the standard protocol. The SBs and FPG-sensitive sites were measured in the same experiment, indicating that the large spread in the latter lesions was the main reason for the reduced inter-laboratory variation. However, it remains worrying that half of the participating laboratories obtained poor results using the standard procedure. This study indicates that future comet assay validation trials should take steps to evaluate the implementation of standard procedures in participating laboratories. This work was supported by ECNIS (Environmental Cancer Risk, Nutrition and Individual Susceptibility), a network of excellence operating within the European Union 6th Framework Program, Priority 5: “Food Quality and Safety” (contract no. 513943).
- Published
- 2012
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