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Inter-laboratory variation in measurement of DNA damage by the alkaline comet assay in the hCOMET ring trial

2023-05-25Journal article Open access
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dc.contributor.authorMøller, Peter
dc.contributor.authorAzqueta, Amaya
dc.contributor.authorCollia, Miguel
dc.contributor.authorBakuradze, Tamara
dc.contributor.authorRichling, Elke
dc.contributor.authorBankoglu, Ezgi Eyluel
dc.contributor.authorStopper, Helga
dc.contributor.authorBastos, Victoria Claudino
dc.contributor.authorLangie, Sabine A.S.
dc.contributor.authorJensen, Annie
dc.contributor.authorRistori, Sara
dc.contributor.authorScavone, Francesca
dc.contributor.authorGiovannelli, Lisa
dc.contributor.authorWojewódzka, Maria
dc.contributor.authorKruszewski, Marcin
dc.contributor.authorValdiglesias, Vanessa
dc.contributor.authorLaffon, Blanca
dc.contributor.authorCosta, Carla
dc.contributor.authorCosta, Solange
dc.contributor.authorTeixeira, João Paulo
dc.contributor.authorMarino, Mirko
dc.contributor.authorDel Bo, Cristian
dc.contributor.authorRiso, Patrizia
dc.contributor.authorZheng, Congying
dc.contributor.authorShaposhnikov, Sergey
dc.contributor.authorCollins, Andrew
dc.date.accessioned2024-02-08T15:49:54Z
dc.date.available2024-02-08T15:49:54Z
dc.date.issued2023-05-25
dc.description.abstractThe comet assay is a simple and versatile method for measurement of DNA damage in eukaryotic cells. More specifically, the assay detects DNA migration from agarose gel-embedded nucleoids, which depends on assay conditions and the level of DNA damage. Certain steps in the comet assay procedure have substantial impact on the magnitude of DNA migration (e.g. electric potential and time of electrophoresis). Inter-laboratory variation in DNA migration levels occurs because there is no agreement on optimal assay conditions or suitable assay controls. The purpose of the hCOMET ring trial was to test potassium bromate (KBrO3) as a positive control for the formamidopyrimidine DNA glycosylase (Fpg)-modified comet assay. To this end, participating laboratories used semi-standardized protocols for cell culture (i.e. cell culture, KBrO3 exposure, and cryopreservation of cells) and comet assay procedures, whereas the data acquisition was not standardized (i.e. staining of comets and image analysis). Segregation of the total variation into partial standard deviation (SD) in % Tail DNA units indicates the importance of cell culture procedures (SD = 10.9), comet assay procedures (SD = 12.3), staining (SD = 7.9) and image analysis (SD = 0.5) on the overall inter-laboratory variation of DNA migration (SD = 18.2). Future studies should assess sources of variation in each of these steps. On the positive side, the hCOMET ring trial demonstrates that KBrO3 is a robust positive control for the Fpg-modified comet assay. In conclusion, the hCOMET ring trial has demonstrated a high reproducibility of detecting genotoxic effects by the comet assay, but inter-laboratory variation of DNA migration levels is a concern.pt_PT
dc.description.sponsorshipCertain authors have acknowledged funding as follows: statutory funding for INTC and IRH (Maria Wojewódzka and Marcin Kruszewski); Spanish Ministry of Science and Innovation (Ref.: PID2020-115348RB-I00) (Amaya Azqueta and Miguel Collia); Spanish Ministry of Science and Innovation MCIN/AEI/10.13039/501100011033 (Grants PID2020- 113788RB-I00 and PID2020-114908 GA-I00), and Xunta de Galicia (ED431B 2022/16) (Vanessa Valdiglesias and Blanca Laffon); Università degli Studi di Firenze Fund RICATEN21 and RICATEN22 (Lisa Giovannelli).pt_PT
dc.description.versioninfo:eu-repo/semantics/publishedVersionpt_PT
dc.identifier.citationMutagenesis. 2023 May 25;38(5):283-294. doi: 10.1093/mutage/gead014pt_PT
dc.identifier.doi10.1093/mutage/gead014pt_PT
dc.identifier.issn0267-8357
dc.identifier.urihttp://hdl.handle.net/10400.18/9058
dc.language.isoengpt_PT
dc.peerreviewedyespt_PT
dc.publisherOxford University Press/ UK Environmental Mutagen Society.pt_PT
dc.relation.publisherversionhttps://academic.oup.com/mutage/article/38/5/283/7179820?login=truept_PT
dc.rights.urihttp://creativecommons.org/licenses/by-nc/4.0/pt_PT
dc.subjectComet Assaypt_PT
dc.subjectDNA Damagept_PT
dc.subjectInter-laboratory Variationpt_PT
dc.subjectRing Trialpt_PT
dc.subjectValidationpt_PT
dc.subjectGenotoxicidade Ambientalpt_PT
dc.titleInter-laboratory variation in measurement of DNA damage by the alkaline comet assay in the hCOMET ring trialpt_PT
dc.typejournal article
dspace.entity.typePublication
oaire.citation.endPage294pt_PT
oaire.citation.issue5pt_PT
oaire.citation.startPage283pt_PT
oaire.citation.titleMutagenesispt_PT
oaire.citation.volume38pt_PT
rcaap.embargofctAcesso de acordo com política editorial da revista.pt_PT
rcaap.rightsopenAccesspt_PT
rcaap.typearticlept_PT

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