An ECVAG inter-laboratory validation study of the comet assay: inter-laboratory and intra-laboratory variations of DNA strand breaks and FPG-sensitive sites in human mononuclear cells.

2.50
Hdl Handle:
http://hdl.handle.net/10146/282455
Title:
An ECVAG inter-laboratory validation study of the comet assay: inter-laboratory and intra-laboratory variations of DNA strand breaks and FPG-sensitive sites in human mononuclear cells.
Authors:
Ersson, Clara; Møller, Peter; Forchhammer, Lykke; Loft, Steffen; Azqueta, Amaya; Godschalk, Roger W. L.; van Schooten, Frederik-Jan; Jones, George D. D.; Higgins, Jennifer A.; Cooke, Marcus S.; Mistry, Vilas; Karbaschi, Mahsa; Phillips, David H.; Sozeri, Osman; Routledge, Michael N.; Nelson-Smith, Kirsty; Riso, Patrizia; Porrini, Marisa; Matullo, Giuseppe; Allione, Alessandra; Stepnik, Maciej; Ferlinska, Magdalena; Teixeira, João Paulo; Costa, Solange; Corcuera, Laura-Ana; López de Cerain, Adela; Laffon, Blanca; Valdiglesias, Vanessa; Collins, Andrew R.; Möller, Lennart
Abstract:
The alkaline comet assay is an established, sensitive method extensively used in biomonitoring studies. This method can be modified to measure a range of different types of DNA damage. However, considerable differences in the protocols used by different research groups affect the inter-laboratory comparisons of results. The aim of this study was to assess the inter-laboratory, intra-laboratory, sample and residual (unexplained) variations in DNA strand breaks and formamidopyrimidine DNA glycosylase (FPG)-sensitive sites measured by the comet assay by using a balanced Latin square design. Fourteen participating laboratories used their own comet assay protocols to measure the level of DNA strand breaks and FPG-sensitive sites in coded samples containing peripheral blood mononuclear cells (PBMC) and the level of DNA strand breaks in coded calibration curve samples (cells exposed to different doses of ionising radiation) on three different days of analysis. Eleven laboratories found dose-response relationships in the coded calibration curve samples on two or three days of analysis, whereas three laboratories had technical problems in their assay. In the coded calibration curve samples, the dose of ionising radiation, inter-laboratory variation, intra-laboratory variation and residual variation contributed to 60.9, 19.4, 0.1 and 19.5%, respectively, of the total variation. In the coded PBMC samples, the inter-laboratory variation explained the largest fraction of the overall variation of DNA strand breaks (79.2%) and the residual variation (19.9%) was much larger than the intra-laboratory (0.3%) and inter-subject (0.5%) variation. The same partitioning of the overall variation of FPG-sensitive sites in the PBMC samples indicated that the inter-laboratory variation was the strongest contributor (56.7%), whereas the residual (42.9%), intra-laboratory (0.2%) and inter-subject (0.3%) variations again contributed less to the overall variation. The results suggest that the variation in DNA damage, measured by comet assay, in PBMC from healthy subjects is assay variation rather than variation between subjects.
Citation:
Mutagenesis 2013, 28 (3):279-286
Journal:
Mutagenesis
Issue Date:
27-Feb-2013
URI:
http://hdl.handle.net/10146/282455
DOI:
10.1093/mutage/get001
PubMed ID:
23446176
Additional Links:
http://mutage.oxfordjournals.org/content/early/2013/02/22/mutage.get001.long
Type:
Article
Language:
en
ISSN:
1464-3804
Sponsors:
This work was supported by Environmental Cancer Risk, Nutrition and Individual Susceptibility (ECNIS), a network of excellence operating within the European Union 6th Framework Program, Priority 5: ‘Food Quality and Safety’ (Contract No 513943), the Swedish Research Council for Environment, Agricultural Sciences and Spatial Planning (FORMAS) and the Swedish Research Council (Vetenskapsrådet). ECVAG (the European Comet assay Validation Group) was created within the ECNIS (Environmental Cancer Risk, Nutrition and Individual Susceptibility) network of excellence, in order to validate the comet assay.
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Full metadata record

DC FieldValue Language
dc.contributor.authorErsson, Claraen_GB
dc.contributor.authorMøller, Peteren_GB
dc.contributor.authorForchhammer, Lykkeen_GB
dc.contributor.authorLoft, Steffenen_GB
dc.contributor.authorAzqueta, Amayaen_GB
dc.contributor.authorGodschalk, Roger W. L.en_GB
dc.contributor.authorvan Schooten, Frederik-Janen_GB
dc.contributor.authorJones, George D. D.en_GB
dc.contributor.authorHiggins, Jennifer A.en_GB
dc.contributor.authorCooke, Marcus S.en_GB
dc.contributor.authorMistry, Vilasen_GB
dc.contributor.authorKarbaschi, Mahsaen_GB
dc.contributor.authorPhillips, David H.en_GB
dc.contributor.authorSozeri, Osmanen_GB
dc.contributor.authorRoutledge, Michael N.en_GB
dc.contributor.authorNelson-Smith, Kirstyen_GB
dc.contributor.authorRiso, Patriziaen_GB
dc.contributor.authorPorrini, Marisaen_GB
dc.contributor.authorMatullo, Giuseppeen_GB
dc.contributor.authorAllione, Alessandraen_GB
dc.contributor.authorStepnik, Maciejen_GB
dc.contributor.authorFerlinska, Magdalenaen_GB
dc.contributor.authorTeixeira, João Pauloen_GB
dc.contributor.authorCosta, Solangeen_GB
dc.contributor.authorCorcuera, Laura-Anaen_GB
dc.contributor.authorLópez de Cerain, Adelaen_GB
dc.contributor.authorLaffon, Blancaen_GB
dc.contributor.authorValdiglesias, Vanessaen_GB
dc.contributor.authorCollins, Andrew R.en_GB
dc.contributor.authorMöller, Lennarten_GB
dc.date.accessioned2013-04-18T09:58:11Z-
dc.date.available2013-04-18T09:58:11Z-
dc.date.issued2013-02-27-
dc.identifier.citationMutagenesis 2013, 28 (3):279-286en_GB
dc.identifier.issn1464-3804-
dc.identifier.pmid23446176-
dc.identifier.doi10.1093/mutage/get001-
dc.identifier.urihttp://hdl.handle.net/10146/282455-
dc.description.abstractThe alkaline comet assay is an established, sensitive method extensively used in biomonitoring studies. This method can be modified to measure a range of different types of DNA damage. However, considerable differences in the protocols used by different research groups affect the inter-laboratory comparisons of results. The aim of this study was to assess the inter-laboratory, intra-laboratory, sample and residual (unexplained) variations in DNA strand breaks and formamidopyrimidine DNA glycosylase (FPG)-sensitive sites measured by the comet assay by using a balanced Latin square design. Fourteen participating laboratories used their own comet assay protocols to measure the level of DNA strand breaks and FPG-sensitive sites in coded samples containing peripheral blood mononuclear cells (PBMC) and the level of DNA strand breaks in coded calibration curve samples (cells exposed to different doses of ionising radiation) on three different days of analysis. Eleven laboratories found dose-response relationships in the coded calibration curve samples on two or three days of analysis, whereas three laboratories had technical problems in their assay. In the coded calibration curve samples, the dose of ionising radiation, inter-laboratory variation, intra-laboratory variation and residual variation contributed to 60.9, 19.4, 0.1 and 19.5%, respectively, of the total variation. In the coded PBMC samples, the inter-laboratory variation explained the largest fraction of the overall variation of DNA strand breaks (79.2%) and the residual variation (19.9%) was much larger than the intra-laboratory (0.3%) and inter-subject (0.5%) variation. The same partitioning of the overall variation of FPG-sensitive sites in the PBMC samples indicated that the inter-laboratory variation was the strongest contributor (56.7%), whereas the residual (42.9%), intra-laboratory (0.2%) and inter-subject (0.3%) variations again contributed less to the overall variation. The results suggest that the variation in DNA damage, measured by comet assay, in PBMC from healthy subjects is assay variation rather than variation between subjects.en_GB
dc.description.sponsorshipThis work was supported by Environmental Cancer Risk, Nutrition and Individual Susceptibility (ECNIS), a network of excellence operating within the European Union 6th Framework Program, Priority 5: ‘Food Quality and Safety’ (Contract No 513943), the Swedish Research Council for Environment, Agricultural Sciences and Spatial Planning (FORMAS) and the Swedish Research Council (Vetenskapsrådet). ECVAG (the European Comet assay Validation Group) was created within the ECNIS (Environmental Cancer Risk, Nutrition and Individual Susceptibility) network of excellence, in order to validate the comet assay.en_GB
dc.languageENG-
dc.language.isoenen
dc.relation.urlhttp://mutage.oxfordjournals.org/content/early/2013/02/22/mutage.get001.longen_GB
dc.rightsArchived with thanks to Mutagenesisen_GB
dc.subjectDNA damageen_GB
dc.subjectComet assayen_GB
dc.subjectHumanen_GB
dc.subjectInter-laboratory validationen_GB
dc.subjectLaboratoriesen_GB
dc.subject.meshAdulten
dc.subject.meshComet Assayen
dc.subject.meshDNA Breaksen
dc.subject.meshDNA-Formamidopyrimidine Glycosylaseen
dc.subject.meshDose-Response Relationshipen
dc.subject.meshFemaleen
dc.subject.meshGamma Raysen
dc.subject.meshHumansen
dc.subject.meshLeukocytes, Mononuclearen
dc.subject.meshMiddle Ageden
dc.subject.meshReproducibility of Resultsen
dc.titleAn ECVAG inter-laboratory validation study of the comet assay: inter-laboratory and intra-laboratory variations of DNA strand breaks and FPG-sensitive sites in human mononuclear cells.en
dc.typeArticleen
dc.identifier.journalMutagenesisen_GB

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