Cellular repair enzymes remove virtually all DNA damage before it is fixed; repair therefore plays a crucial role in preventing cancer. Repair studied at the level of transcription correlates poorly with enzyme activity, and so assays of phenotype are needed. In a biochemical approach, substrate nucleoids containing specific DNA lesions are incubated with cell extract; repair enzymes in the extract induce breaks at damage sites; and the breaks are measured with the comet assay. The nature of the substrate lesions defines the repair pathway to be studied. This in vitro DNA repair assay has been modified for use in animal tissues, specifically to study the effects of aging and nutritional intervention on repair. Recently, the assay was applied to different strains of Drosophila melanogaster proficient and deficient in DNA repair. Most applications of the repair assay have been in human biomonitoring. Individual DNA repair activity may be a marker of cancer susceptibility; alternatively, high repair activity may result from induction of repair enzymes by exposure to DNA-damaging agents. Studies to date have examined effects of environment, nutrition, lifestyle, and occupation, in addition to clinical investigations.

Department of Genetics and Biotechnology Animal and Veterinary Research Centre University of Trás os Montes and Alto Douro Vila Real Portugal

Department of Molecular Biology of Cancer Institute of Experimental Medicine Academy of Science of the Czech Republic Prague Czech Republic

Department of Nutrition University of Oslo Oslo Norway

Department of Pharmacology and Toxicology Faculty of Pharmacy University of Navarra Pamplona Spain

Environmental Risk and Health Unit Flemish Institute of Technological Research Mol Belgium

Zobrazit více v PubMed

Adams M. D., Celniker S. E., Holt R. A., Evans C. A., Gocayne J. D., Amanatides P. G., et al. (2000). The genome sequence of Drosophila melanogaster. Science 287 2185–2195 10.1126/science.287.5461.2185 PubMed DOI

Au W. W., Giri A. A., Ruchirawat M. (2010). Challenge assay: a functional biomarker for exposure-induced DNA repair deficiency and for risk of cancer. Int. J. Hyg. Environ. Health 213 32–39 10.1016/j.ijheh.2009.09.002 PubMed DOI

Azqueta A., Collins A. R. (2013). The essential comet assay: a comprehensive guide to measuring DNA damage and repair. Arch. Toxicol. 87 949–968 10.1007/s00204-013-1070-0 PubMed DOI

Azqueta A., Langie S. A. S., Slyskova J., Collins A. R. (2013a). Measurement of DNA base and nucleotide excision repair activities in mammalian cells and tissues using the comet assay – a methodological overview. DNA Repair 12 1007–1010 10.1016/j.dnarep.2013.07.011 PubMed DOI

Azqueta A., Costa S., Lorenzo Y., Bastani N. E., Collins A. R. (2013b). Vitamin C in cultured human (HeLa) cells: lack of effect on DNA protection and repair. Nutrients 5 1200–1217 10.3390/nu5041200 PubMed DOI PMC

Azqueta A., Lorenzo Y., Collins A. R. (2009). In vitro comet assay for DNA repair: a warning concerning application to cultured cells. Mutagenesis 24 379–381 10.1093/mutage/gep009 PubMed DOI

Brevik A., Gaivão I., Medin T., Jørgensen A., Piasek A., Elliasson J., et al. (2011a). Supplementation of a western diet with golden kiwifruits (Actinidia chinensis var. ‘Hort 16A’): effects on biomarkers of oxidation damage and antioxidant protection. Nutr. J. 10 54 10.1186/1475-2891-10-54 PubMed DOI PMC

Brevik A., Karlsen A., Azqueta A., Estaban A. T., Blomhoff R., Collins A. R. (2011b). Both base excision repair and nucleotide excision repair in humans are influenced by nutritional factors. Cell Biochem. Funct. 29 36–42 10.1002/cbf.1715 PubMed DOI

Caple F., Williams E. A., Spiers A., Tyson J., Burtle B., Daly A. K., et al. (2010). Inter-individual variation in DNA damage and base excision repair in young, healthy non-smokers: effects of dietary supplementation and genotype. Br. J. Nutr. 103 1585–1593 10.1017/S0007114509993540 PubMed DOI

Collins A. R., Azqueta A. (2012). DNA repair as a biomarker in human biomonitoring studies: further applications of the comet assay. Mutat. Res. 736 122–129 10.1016/j.mrfmmm.2011.03.005 PubMed DOI

Collins A. R., Azqueta A., Langie S. A. S. (2012). Effects of micronutrients on DNA repair. Eur. J. Nutr. 51 261–279 10.1007/s00394-012-0318-4 PubMed DOI

Collins A. R., Dusinska M., Horvathova E., Munro E., Savio M., Stetina R. (2001). Inter-individual differences in repair of DNA base oxidation, measured in vitro with the comet assay. Mutagenesis 16 297–301 10.1093/mutage/16.4.297 PubMed DOI

Collins A. R., Fleming I. M., Gedik C. M. (1994). In vitro repair of oxidative and ultraviolet-induced DNA damage in supercoiled nucleoid DNA by human cell extract. Biochim. Biophys. Acta 1219 724–727 10.1016/0167-4781(94)90236-4 PubMed DOI

Collins A. R., Harrington V., Drew J., Melvin R. (2003). Nutritional modulation of DNA repair in a human intervention study. Carcinogenesis 24 511–515 10.1093/carcin/24.3.511 PubMed DOI

Cook P. R., Brazell I. A., Jost E. (1976). Characterization of nuclear structures containing superhelical DNA. J. Cell Sci. 22 303–324 PubMed

Dusinska M., Collins A. R., Kazimirova A., Barancokova M., Harrington V., Volkovova K., et al. (2004a). Genotoxic effects of asbestos in humans. Mutat. Res. 553 91–102 10.1016/j.mrfmmm.2004.06.027 PubMed DOI

Dusinska M., Barancokova M., Kazimirova A., Harrington V., Volkovova K., Staruchova M., et al. (2004b). Does occupational exposure to mineral fibres cause DNA or chromosome damage? Mutat. Res. 553 103–110 10.1016/j.mrfmmm.2004.06.029 PubMed DOI

Etemadi A., Islami F., Phillips D. H., Godschalk R., Golozar A., Kamangar F., et al. (2013). Variation in PAH-related DNA adduct levels among non-smokers: the role of multiple genetic polymorphisms and nucleotide excision repair phenotype. Int. J. Cancer 132 2738–2747 10.1002/ijc.27953 PubMed DOI PMC

Friedberg E. C., Walker G. C., Siede W., Wood R. D., Schultz R. A., Ellenberger T. (2006). “Correcting altered bases in DNA: DNA repair,” in DNA Repair and Mutagenesis Pt 2 eds Friedberg E. C., Walker G. C., Siede W., Wood R. D., Schultz R. A., Ellenberger T. (Washington, DC: ASM Press; ) 107–460

Gaivão I., Piasek A., Brevik A., Shaposhnikov S., Collins A. R. (2009). Comet assay-based methods for measuring DNA repair in vitro; estimates of inter- and intra-individual variation. Cell Biol. Toxicol. 25 45–52 10.1007/s10565-007-9047-5 PubMed DOI

Gaivão I., Rodriguez R., Sierra L. M. (2014). “Use of the comet assay to study DNA repair in Drosophila melanogaster,” in Genotoxicity and DNA Repair: A Practical Approach (Series: Methods in Pharmacology and Toxicology) eds Sierra L. M., Gaivão I. (New York, NY: Humana Press; ).

Gedik C. M., Ewen S. W., Collins A. R. (1992). Single-cell gel electrophoresis applied to the analysis of UV-C damage and its repair in human cells. Int. J. Radiat. Biol. 62 313–320 10.1080/09553009214552161 PubMed DOI

Gorniak J. P., Cameron K. M., Waldron K. J., von Zglinicki T., Mathers J. C., Langie S. A. (2013). Tissue differences in BER-related incision activity and non-specific nuclease activity as measured by the comet assay. Mutagenesis 28 673–681 10.1093/mutage/get047 PubMed DOI

Guarnieri S., Loft S., Riso P., Porrini M., Risom L., Poulsen H., et al. (2008). DNA repair phenotype and dietary antioxidant supplementation. Br. J. Nutr. 99 1018–1024 10.1017/S0007114507842796 PubMed DOI

Guo J., Hanawalt P. C., Spivak G.(2013). Comet-FISH with strand-specific probes reveals transcription-coupled repair of 8-oxoGuanine in human cells. Nucleic Acids Res. 41 7700–7712 10.1093/nar/gkt524 PubMed DOI PMC

Hallström I., Blank A., Atuma S. (1984). Genetic variation in cytochrome P450 and xenobiotic metabolism in Drosophila melanogaster. Biochem. Pharmacol. 33 13–20 10.1016/0006-2952(84)90364-2 PubMed DOI

Hasplova K., Hudecova A., Magdolenova Z., Bjoras M., Galova E., Miadokova E., et al. (2012). DNA alkylation lesions and their repair in human cells: modification of the comet assay with 3-methyladenine DNA glycosylase (AlkD). Toxicol. Lett. 208 76–81 10.1016/j.toxlet.2011.10.005 PubMed DOI

Herrera M., Dominguez G., Garcia J. M., Pena C., Jimenez C., Silva J., et al. (2009). Differences in repair of DNA cross-links between lymphocytes and epithelial tumor cells from colon cancer patients measured in vitro with the comet assay. Clin. Cancer Res. 15 5466–5472 10.1158/1078-0432.CCR-08-3268 PubMed DOI

Langie S. A., Achterfeldt S., Gorniak J. P., Halley-Hogg K. J., Oxley D., van Schooten F. J., et al. (2013). Maternal folate depletion and high-fat feeding from weaning affects DNA methylation and DNA repair in brain of adult offspring. FASEB J. 27 3323–3334 10.1096/fj.12-224121 PubMed DOI

Langie S. A., Cameron K. M., Waldron K. J., Fletcher K. P., von Zglinicki T., Mathers J. C. (2011). Measuring DNA repair incision activity of mouse tissue extracts towards singlet oxygen-induced DNA damage: a comet-based in vitro repair assay. Mutagenesis 26 461–471 10.1093/mutage/ger005 PubMed DOI

Langie S. A., Kowalczyk P., Tudek B., Zabielski R., Dziaman T., Olinski R., et al. (2010a). The effect of oxidative stress on nucleotide-excision repair in colon tissue of newborn piglets. Mutat. Res. 695 75–80 10.1016/j.mrgentox.2009.12.005 PubMed DOI

Langie S. A. S., Wilms L. C., Hämäläinen S., Kleinjans J. C. S., Godschalk R. W. L., van Schooten F. J. (2010b). Modulation of nucleotide excision repair in human lymphocytes by genetic and dietary factors. Br. J. Nutr. 103 490–501 10.1017/S0007114509992066 PubMed DOI

Langie S. A. S., Knaapen A. M., Brauers K. J. J., van Berlo D., van Schooten F.-J., Godschalk R. W. L. (2006). Development and validation of a modified comet assay to phenotypically assess nucleotide excision repair. Mutagenesis 21 153–158 10.1093/mutage/gel013 PubMed DOI

Langie S. A. S., Kowalczyk P., Tomaszewski B., Vasilaki A., Maas L. M., Moonen E. J., et al. (2014). Redox and epigenetic regulation of the APE1 gene in the hippocampus of piglets: the effect of early life exposures. DNA Repair 18 52–62 10.1016/j.dnarep.2014.03.011 PubMed DOI

Lorenzo Y., Azqueta A., Luna L., Bonilla F., Dominguez G., Collins A. R. (2009). The carotenoid β-cryptoxanthin stimulates the repair of DNA oxidation damage in addition to acting as an antioxidant in human cells. Carcinogenesis 30 308–314 10.1093/carcin/bgn270 PubMed DOI

Mikkelsen L., Bialkowski K., Risom L., Løhr M., Loft S., Møller P. (2009). Aging and defense against generation of 8-oxo-8-dihydro-2′-deoxyguanosine in DNA. Free Radic. Biol. Med. 47 608–615 10.1016/j.freeradbiomed.2009.05.030 PubMed DOI

Ostling O., Johanson K. J. (1984). Microelectrophoretic study of radiation-induced DNA damages in individual mammalian cells. Biochem. Biophys. Res. Commun. 123 291–298 10.1016/0006-291X(84)90411-X PubMed DOI

Paz-Elizur T., Elinger D., Leitner-Dagan Y., Blumenstein S., Krupsky M., Berrebi A., et al. (2007). Development of an enzymatic DNA repair assay for molecular epidemiology studies: distribution of OGG activity in healthy individuals. DNA Repair 6 45–60 10.1016/j.dnarep.2006.08.003 PubMed DOI

Ramos A. A., Azqueta A., Pereira-Wilson C., Collins A. R. (2010a). Polyphenolic compounds from Salvia species protect cellular DNA from oxidation and stimulate DNA repair in cultured human cells. J. Agric. Food Chem. 58 7465–7471 10.1021/jf100082p PubMed DOI

Ramos A. A., Pereira-Wilson C., Collins A. R. (2010b). Protective effects of ursolic acid and luteolin against oxidative DNA damage include enhancement of DNA repair in Caco-2 cells. Mutat. Res. 692 6–11 10.1016/j.mrfmmm.2010.07.004 PubMed DOI

Riso P., Martini D., Møller P., Loft S., Bonacina G., Moro M., et al. (2010). DNA damage and repair activity after broccoli intake in young healthy smokers. Mutagenesis 25 595–602 10.1093/mutage/geq045 PubMed DOI

Sekelsky J. J., Brodsky M. H., Burtis K. C. (2000). DNA repair in Drosophila: insights from the Drosophila genome sequence. J. Cell. Biol. 150 31–36 10.1083/jcb.150.2.F31 PubMed DOI PMC

Shaposhnikov S., Thomsen P. D., Collins A. R. (2011). “Combining fluorescent in situ hybridization with the comet assay for targeted examination of DNA damage and repair. Methods Mol. Biol. 682 115–132 10.1007/978-1-60327-409-8_10 PubMed DOI

Silva J. P., Gomes A. C., Coutinho O. P. (2008). Oxidative DNA damage protection and repair by polyphenolic compounds in PC12 cells. Eur. J. Pharmacol. 601 50–56 10.1016/j.ejphar.2008.10.046 PubMed DOI

Silva J. P., Gomes A. C., Proenca F., Coutinho O. P. (2009). Novel nitrogen compounds enhance protection and repair of oxidative DNA damage in a neuronal cell model: comparison with quercetin. Chem. Biol. Interact. 181 328–337 10.1016/j.cbi.2009.07.024 PubMed DOI

Singh N. P., McCoy M. T., Tice R. R., Schneider E. L. (1988). A simple technique for quantitation of low levels of DNA damage in individual cells. Exp. Cell Res. 175 184–191 10.1016/0014-4827(88)90265-0 PubMed DOI

Sliwinski T., Czechowska A., Szemraj J., Morawiec Z., Skorski T., Blasiak J. (2008). STI571 reduces NER activity in BCR/ABL-expressing cells. Mutat. Res. 654 162–167 10.1016/j.mrgentox.2008.06.002 PubMed DOI

Slyskova J., Dusinska M., Kuricova M., Soucek P., Vodickova L., Susova S., et al. (2007). Relationship between the capacity to repair 8-oxoguanine, biomarkers of genotoxicity and individual susceptibility in styrene-exposed workers. Mutat. Res. 634 101–111 10.1016/j.mrgentox.2007.06.012 PubMed DOI

Slyskova J., Korenkova V., Collins A. R., Prochazka P., Vodickova L., Svec J., et al. (2012). Functional, genetic, and epigenetic aspects of base and nucleotide excision repair in colorectal carcinomas. Clin. Cancer Res. 18 5878–5887 10.1158/1078-0432.CCR-12-1380 PubMed DOI

Slyskova J., Lorenzo Y., Karlsen A., Carlsen M. H., Novosadova V., Blomhoff R., et al. (2014a). Both genetic and dietary factors underlie individual differences in DNA damage levels and DNA repair capacity. DNA Repair. 16 66–73 10.1016/j.dnarep.2014.01.016 PubMed DOI

Slyskova J., Langie S. A. S., Collins A. R., Vodicka P. (2014b). Functional evaluation of DNA repair in human biopsies and their relation to other cellular biomarkers. Front. Genet. 5:116 10.3389/fgene.2014.00116 PubMed DOI PMC

Slyskova J., Langie S. A. S., Gaivão I., Collins A. R., Azqueta A. (2014c). “A standardized protocol for the in vitro comet-based DNA repair assay,” Genotoxicity and DNA Repair: A Practical Approach, Methods in Pharmacol.ogy and Toxicology eds Sierra L. M., Gaivão I. (New York, NY: Humana Press; ) 377–395

Søndergaard L. (1993). Homology between the mammalian liver and the Drosophila fat body. Trends Genet. 9 193 10.1016/0168-9525(93)90113-V PubMed DOI

Spanswick V. J., Hartley J. M., Hartley J. A. (2010). Measurement of DNA interstrand crosslinking in individual cells using the single cell gel electrophoresis (Comet) assay. Methods Mol. Biol. 613 267–282 10.1007/978-1-60327-418-0_17 PubMed DOI

Spivak G., Cox R. A., Hanawalt P. C. (2009). New applications of the Comet assay: comet-FISH and transcription-coupled DNA repair. Mutat. Res. 681 44–50 10.1016/j.mrrev.2007.12.003 PubMed DOI PMC

Stoyanova E., Pastor S., Coll E., Azqueta A., Collins A. R., Marcos R. (2014). Base excision repair capacity in chronic renal failure patients undergoing hemodialysis treatment. Cell Biochem. Funct. 32 177–182 10.1002/cbf.2989 PubMed DOI

Tomasetti M., Alleva R., Borghi B., Collins A. R. (2001). In vivo supplementation with coenzyme Q10 enhances the recovery of human lymphocytes from oxidative DNA damage. FASEB J. 15 1425–1427 10.1096/fj.00-0694fje PubMed DOI

Tsai Y. C., Li P. Y., Chen C. C., Liu Y. C. (2013). Is the oxidative DNA damage level of human lymphocyte correlated with the antioxidant capacity of serum or the base excision repair activity of lymphocyte? Oxid. Med. Cell. Longev. 2013 237583 10.1155/2013/237583 PubMed DOI PMC

Tyson J., Mathers J. C. (2007). Dietary and genetic modulation of DNA repair in healthy human adults. Proc. Nutr. Soc. 66 42–51 10.1017/S0029665107005289 PubMed DOI

Vande Loock K., Decordier I., Ciardelli R., Haumont D., Kirsch-Volders M. (2010). An aphidicolin-block nucleotide excision repair assay measuring DNA incision and repair capacity. Mutagenesis 25 25–32 10.1093/mutage/gep039 PubMed DOI

van Dyk E., Steenkamp A., Koekemoer G., Pretorius P. J. (2010). Hereditary tyrosinemia type 1 metabolites impair DNA excision repair pathways. Biochem. Biophys. Res. Commun. 401 32–36 10.1016/j.bbrc.2010.09.002 PubMed DOI

An optimized comet-based in vitro DNA repair assay to assess base and nucleotide excision repair activity