The comet assay: topical issues
Andrew Collins,Amaia Azqueta Oscoz,Gunnar Brunborg,Isabel Gaivão,Isabel Gaivão,Lisa Giovannelli,Marcin Kruszewski,Catherine C. Smith,Rudolf Stetina +8 more
Reads0
Chats0
TLDR
The comet assay is a versatile and sensitive method for measuring single- and double-strand breaks in DNA, and by including lesion-specific enzymes in the assay, its range and sensitivity are greatly increased, but it is important to bear in mind that their specificity is not absolute.Abstract:
The comet assay is a versatile and sensitive method for measuring single- and double-strand breaks in DNA. The mechanism of formation of comets (under neutral or alkaline conditions) is best understood by analogy with nucleoids, in which relaxation of DNA supercoiling in a structural loop of DNA by a single DNA break releases that loop to extend into a halo-or, in the case of the comet assay, to be pulled towards the anode under the electrophoretic field. A consideration of the simple physics underlying electrophoresis leads to a better understanding of the assay. The sensitivity of the assay is only fully appreciated when it is calibrated: between one hundred and several thousand breaks per cell can be determined. By including lesion-specific enzymes in the assay, its range and sensitivity are greatly increased, but it is important to bear in mind that their specificity is not absolute. Different approaches to quantitation of the comet assay are discussed. Arguments are presented against trying to apply the comet assay to the study of apoptosis. Finally, some of the advantages and disadvantages of using the comet assay on lymphocyte samples collected in human studies are rehearsed.read more
Citations
More filters
Journal ArticleDOI
NanoGenotoxicology: The DNA damaging potential of engineered nanomaterials
Neenu Singh,Bella B. Manshian,Gareth J.S. Jenkins,Sioned M. Griffiths,Paul M. Williams,Thierry G.G. Maffeis,Chris J. Wright,Shareen H. Doak +7 more
TL;DR: Many of the engineered nanomaterials assessed were found to cause genotoxic responses, such as chromosomal fragmentation, DNA strand breakages, point mutations, oxidative DNA adducts and alterations in gene expression profiles.
Journal ArticleDOI
Arsenic-induced oxidative stress and its reversibility
TL;DR: This review summarizes the literature describing the molecular mechanisms of arsenic-induced oxidative stress, its relevant biomarkers, and its relation to various diseases, including preventive and therapeutic strategies and updates the reader on recent advances in chelation therapy and newer therapeutic strategies suggested to treat arsenic- induced oxidative damage.
Journal ArticleDOI
Glyphosate-based herbicides are toxic and endocrine disruptors in human cell lines.
Céline Gasnier,Coralie Dumont,Nora Benachour,Emilie Clair,Marie-Christine Chagnon,Gilles-Eric Séralini +5 more
TL;DR: A real cell impact of glyphosate-based herbicides residues in food, feed or in the environment has thus to be considered, and their classifications as carcinogens/mutagens/reprotoxics is discussed.
Journal ArticleDOI
H2AX Phosphorylation: Its Role in DNA Damage Response and Cancer Therapy
TL;DR: Analysis of γH2AX expression can be used to detect the genotoxic effect of different toxic substances and to predict of tumor cell sensitivity to DNA damaging anticancer agents and toxicity of anticancer treatment toward normal cells.
Journal ArticleDOI
The essential comet assay: a comprehensive guide to measuring DNA damage and repair
Amaya Azqueta,Andrew Collins +1 more
TL;DR: The comet assay has been modified to detect various base alterations, by including digestion of nucleoids with a lesion-specific endonuclease, and modifications to measure cellular antioxidant status and different types of DNA repair.
References
More filters
Journal ArticleDOI
A simple technique for quantitation of low levels of DNA damage in individual cells
TL;DR: Human lymphocytes were exposed to X-irradiation or treated with H2O2 and the extent of DNA migration was measured using a single-cell microgel electrophoresis technique under alkaline conditions and this technique appears to be sensitive and useful for detecting damage and repair in single cells.
Journal ArticleDOI
Microelectrophoretic study of radiation-induced DNA damages in individual mammalian cells.
O. Ostling,K.J. Johanson +1 more
TL;DR: Mammalian cells were after irradiation suspended in melted agarose, and casted on microscope slides, and DNA had migrated towards the anode and this migration was more pronounced in irradiated than in control cells.
Journal ArticleDOI
Direct enzymic detection of endogenous oxidative base damage in human lymphocyte DNA
TL;DR: Using an endonuclease specific for oxidized pyrimidines, in conjunction with the highly sensitive method of single cell gel electrophoresis, significant oxidative damage is detected in untreated, freshly isolated lymphocytes from normal, healthy individuals.
Journal ArticleDOI
Comet assay in human biomonitoring studies: Reliability, validation, and applications
Andrew Collins,Maria Dusinska,M. F. Franklin,Martina Somorovská,Helena Petrovská,Susan J. Duthie,Laurence Fillion,Mihalis I. Panayiotidis,Katarina Raslova,Nicholas J. Vaughan +9 more
TL;DR: The reliability and reproducibility of the comet assay is reported, from the level of comparing results from duplicate gels prepared from the same sample of cells, up to an assessment of the natural intra‐ and interindividual variability in lymphocyte DNA damage measured in groups of normal, healthy human volunteers.
Journal ArticleDOI
Establishing the background level of base oxidation in human lymphocyte DNA: results of an interlaboratory validation study
TL;DR: The ESCODD consortium of mainly European laboratories has attempted to minimize this artifact and to provide standard, reliable protocols for sample preparation and analysis as mentioned in this paper. But the accuracy of low levels of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) in DNA is hampered by the ease with which guanine is oxidized during preparation of DNA for analysis.