Variation in the measurement of DNA damage by comet assay measured by the ECVAG inter-laboratory validation trial.

2.50
Hdl Handle:
http://hdl.handle.net/10146/114952
Title:
Variation in the measurement of DNA damage by comet assay measured by the ECVAG inter-laboratory validation trial.
Authors:
Forchhammer, Lykke; Johansson, Clara; Loft, Steffen; Moller, Lennart; Godschalk, Roger W. L.; Langie, Sabine A. S.; Jones, George D. D.; Kwok, Rachel W. L.; Collins, Andrew R.; Azqueta, Amaya; Phillips, David H.; Sozeri, Osman; Stepnik, Maciej; Palus, Jadwiga; Vogel, Ulla; Wallin, Hakan; Routledge, Michael N.; Handforth, Catherine; Allione, Alessandra; Matullo, Giuseppe; Teixeira, Joao Paulo; Costa, Solange; Riso, Patrizia; Porrini, Marisa; Moller, Peter
Abstract:
The comet assay has become a popular method for the assessment of DNA damage in biomonitoring studies and genetic toxicology. However, few studies have addressed the issue of the noted inter-laboratory variability of DNA damage measured by the comet assay. In this study, 12 laboratories analysed the level of DNA damage in monocyte-derived THP-1 cells by either visual classification or computer-aided image analysis of pre-made slides, coded cryopreserved samples of cells and reference standard cells (calibration curve samples). The reference standard samples were irradiated with ionizing radiation (0-10 Gy) and used to construct a calibration curve to calculate the number of lesions per 10(6) base pair. All laboratories detected dose-response relationships in the coded samples irradiated with ionizing radiation (1.5-7 Gy), but there were overt differences in the level of DNA damage reported by the different laboratories as evidenced by an inter-laboratory coefficient of variation (CV) of 47%. Adjustment of the primary comet assay end points by a calibration curve prepared in each laboratory reduced the CV to 28%, a statistically significant reduction (P < 0.05, Levene's test). A large fraction of the inter-laboratory variation originated from differences in image analysis, whereas the intra-laboratory variation was considerably smaller than the variation between laboratories. In summary, adjustment of primary comet assay results by reference standards reduces inter-laboratory variation in the level of DNA damage measured by the alkaline version of the comet assay.
Citation:
Mutagenesis 2010, 25 (2):113-123
Journal:
Mutagenesis
Issue Date:
Mar-2010
URI:
http://hdl.handle.net/10146/114952
DOI:
10.1093/mutage/gep048
PubMed ID:
19910383
Additional Links:
http://mutage.oxfordjournals.org/content/25/2/113.long
Type:
Article
Language:
en
ISSN:
1464-3804
Sponsors:
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); the Danish Research Council (to L.F., S.L. and P.M.); the Italian Association for Cancer Research (to A.A. and G.M.); FP6 COMICS Project (contract no. 037575 to A.R.C. and A.A.).
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Full metadata record

DC FieldValue Language
dc.contributor.authorForchhammer, Lykkeen
dc.contributor.authorJohansson, Claraen
dc.contributor.authorLoft, Steffenen
dc.contributor.authorMoller, Lennarten
dc.contributor.authorGodschalk, Roger W. L.en
dc.contributor.authorLangie, Sabine A. S.en
dc.contributor.authorJones, George D. D.en
dc.contributor.authorKwok, Rachel W. L.en
dc.contributor.authorCollins, Andrew R.en
dc.contributor.authorAzqueta, Amayaen
dc.contributor.authorPhillips, David H.en
dc.contributor.authorSozeri, Osmanen
dc.contributor.authorStepnik, Maciejen
dc.contributor.authorPalus, Jadwigaen
dc.contributor.authorVogel, Ullaen
dc.contributor.authorWallin, Hakanen
dc.contributor.authorRoutledge, Michael N.en
dc.contributor.authorHandforth, Catherineen
dc.contributor.authorAllione, Alessandraen
dc.contributor.authorMatullo, Giuseppeen
dc.contributor.authorTeixeira, Joao Pauloen
dc.contributor.authorCosta, Solangeen
dc.contributor.authorRiso, Patriziaen
dc.contributor.authorPorrini, Marisaen
dc.contributor.authorMoller, Peteren
dc.date.accessioned2010-11-08T12:37:57Z-
dc.date.available2010-11-08T12:37:57Z-
dc.date.issued2010-03-
dc.identifier.citationMutagenesis 2010, 25 (2):113-123en
dc.identifier.issn1464-3804-
dc.identifier.pmid19910383-
dc.identifier.doi10.1093/mutage/gep048-
dc.identifier.urihttp://hdl.handle.net/10146/114952-
dc.description.abstractThe comet assay has become a popular method for the assessment of DNA damage in biomonitoring studies and genetic toxicology. However, few studies have addressed the issue of the noted inter-laboratory variability of DNA damage measured by the comet assay. In this study, 12 laboratories analysed the level of DNA damage in monocyte-derived THP-1 cells by either visual classification or computer-aided image analysis of pre-made slides, coded cryopreserved samples of cells and reference standard cells (calibration curve samples). The reference standard samples were irradiated with ionizing radiation (0-10 Gy) and used to construct a calibration curve to calculate the number of lesions per 10(6) base pair. All laboratories detected dose-response relationships in the coded samples irradiated with ionizing radiation (1.5-7 Gy), but there were overt differences in the level of DNA damage reported by the different laboratories as evidenced by an inter-laboratory coefficient of variation (CV) of 47%. Adjustment of the primary comet assay end points by a calibration curve prepared in each laboratory reduced the CV to 28%, a statistically significant reduction (P < 0.05, Levene's test). A large fraction of the inter-laboratory variation originated from differences in image analysis, whereas the intra-laboratory variation was considerably smaller than the variation between laboratories. In summary, adjustment of primary comet assay results by reference standards reduces inter-laboratory variation in the level of DNA damage measured by the alkaline version of the comet assay.en
dc.description.sponsorshipEnvironmental 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); the Danish Research Council (to L.F., S.L. and P.M.); the Italian Association for Cancer Research (to A.A. and G.M.); FP6 COMICS Project (contract no. 037575 to A.R.C. and A.A.).en
dc.language.isoenen
dc.relation.urlhttp://mutage.oxfordjournals.org/content/25/2/113.longen
dc.subjectRadiation, Ionizingen
dc.subjectDNA Damageen
dc.subjectCells, Cultureden
dc.subjectHumansen
dc.subjectComet Assayen
dc.subjectReference Standardsen
dc.subjectLaboratoriesen
dc.subject.meshCalibration-
dc.subject.meshCells, Cultured-
dc.subject.meshComet Assay-
dc.subject.meshCryopreservation-
dc.subject.meshDNA Damage-
dc.subject.meshDNA-Formamidopyrimidine Glycosylase-
dc.subject.meshHumans-
dc.subject.meshLaboratories-
dc.subject.meshMonocytes-
dc.subject.meshObserver Variation-
dc.subject.meshRadiation, Ionizing-
dc.subject.meshReference Standards-
dc.titleVariation in the measurement of DNA damage by comet assay measured by the ECVAG inter-laboratory validation trial.en
dc.typeArticleen
dc.identifier.journalMutagenesisen

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