Using the comet assay to study DNA repair: progress in the past decade.
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Flemish Institute of Technological Research (VITO), Environmental Risk and Health unit, Belgium
The comet assay is a versatile and sensitive method for measuring strand breaks in DNA. The first paper on this single-cell gel electrophoresis assay was published in 1984 by Ostling and Johanson (1984). This assay (with and without inclusion of lesion-specific enzymes) is widely used as a biomarker assay in human population studies - primarily to measure DNA damage, but increasingly also to assess the capacity of cells for DNA repair. Ostling and Johanson were also the first to report experiments to measure DNA repair, by simply following the decrease of DNA damage over time after challenging cells with ionising radiation. However, this approach is time-consuming and laborious, and therefore not ideal for biomonitoring studies, which typically require high-throughput processing of many samples. As an alternative approach, the in vitro comet-based repair assay was developed: a cell extract is incubated with a DNA substrate containing specific lesions, and DNA incisions accumulate. The in vitro comet-based repair assay was first devised to measure base excision repair in lymphocytes (Collins et al., 2001). However, over the past decade it has been modified and improved to study incision of other lesions and thus other repair pathways, as well as being applied to tissue samples in addition to cell suspensions. The application of the in vitro repair assay in dietary intervention, environmental biomonitoring and animal studies will be discussed.
References
Collins, A.R., Dusinska, M., Horvathova, E., Munro, E., Savio, M., and Stetina, R. (2001). Inter-individual differences in repair of DNA base oxidation, measured in vitro with the comet assay. Mutagenesis 16, 297–301. doi:10.1093/mutage/16.4.297
Ostling, O., and Johanson, K.J. (1984). Microelectrophoretic study of radiation-induced DNA damages in individual mammalian cells. Biochem Biophys Res Commun 123,291-298. doi:10.1016/0006-291X(84)90411-X
Keywords:
Comet Assay,
DNA Repair,
DNA Damage,
Single cell gel electrophoresis,
Protein extracts
Conference:
ICAW 2015 - 11th International Comet Assay Workshop, Antwerpen, Belgium, 1 Sep - 4 Sep, 2015.
Presentation Type:
Oral Presentation
Topic:
Origin and history of the Comet Assay
Citation:
Langie
S
(2015). Using the comet assay to study DNA repair: progress in the past decade..
Front. Genet.
Conference Abstract:
ICAW 2015 - 11th International Comet Assay Workshop.
doi: 10.3389/conf.fgene.2015.01.00068
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Received:
16 Apr 2015;
Published Online:
23 Jun 2015.
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Correspondence:
Dr. Sabine Langie, Flemish Institute of Technological Research (VITO), Environmental Risk and Health unit, Mol, Please Select…, 2400, Belgium, sabine.langie@vito.be