Comet Assay

An overview of Andor's solutions for Comet Assay

Comet Assay
Comet image courtesy of Marie Vasquez, Helex 3 Inc.

Comet assay? Not a technique in astronomy whereby the solar system and beyond is scanned to assess the risk of the human race following the fate of the dinosaurs! The Comet Assay, also known as "Single Cell Gel (SCG) electrophoresis", is a highly sensitive and efficient means of assessing and quantifying DNA damage and repair at the level of the individual cell, applicable to any eukaryotic cell. Types of DNA damage include single-/double-strand breaks, DNA-drug crosslinking, DNA-protein or DNA-DNA crosslinking and oxidative DNA base damage.The technique is based on fluorescence microscopy and involves embedding nuclei in agarose gel, then electrophoretic separation across an applied electric field. The DNA is stained with a fluorescent dye, and subsequently imaged under an epifluorescence microscope. The resulting structure is reminiscent of a comet, consisting of the nucleus head, and tail of damaged DNA. During electrophoresis, fractured DNA fragments migrate away from the nucleus and the extent of DNA damage can be quantified by measuring the relative % DNA between head and tail. Importantly, the exact class of DNA damage can be examined by careful control over cell preparation conditions. For example, the extent of alkali treatment (pH control) that is used to convert a DNA damage site into a DNA fragment (essential for Comet detection) can discriminate between different types of strand breaks. Sites of DNA base modifications can be converted to DNA fragments by introduction of enzymes that act specifically on such lesions. Furthermore, DNA crosslinking and binding can also be detected by the Comet assay, by determining the extent to which these interactions retard migration of DNA. Since its "impact", Comet has become a core technique within fields or study such as toxicology, apoptosis, DNA repair, aging, cell cycle analysis and free radical biology.

Comet slides are examined under a fluorescent microscope, often by widefield epifluorescence microscopy. Widefield epifluorescence microscopy has developed into a universally accessible technique for study of fluorescently labelled cells and tissues. Over the last few decades, this drive has been accelerated by improvements in fluorescent probes, labelling chemistry, optical instrumentation (such as filters and objectives) and detector technology.

The widefield technique involves flood-illumination of the field of view by a wavelength or small wavelength range (often though use of an excitation filter and arc lamp). The stoke-shifted fluorescent emission transmits though the dichroic, that was initially used to reflect the shorter wavelength excitation light onto the sample, gets optically filtered once again by an emission filter (often called barrier filter), and focused onto the camera detector.

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Publications Database
Evaluation of genetic damage in tobacco and arsenic exposed population of Southern Assam, India using buccal cytome assay and comet assay
Chloroquine-induced glioma cells death is associated with mitochondrial membrane potential loss, but not oxidative stress
CDC42 GTPase Activation Affects HeLa Cell DNA Repair and Proliferation Following UV Radiation‐Induced Genotoxic Stress
TGFβ1 Protects cells from γ-IR by enhancing the activity of the NHEJ repair pathway
Two-Step Procedure for Evaluating Experimentally Induced DNA Damage: Texas Red and Comet Assays
Activation of DNA damage repair pathways in response to nitrogen mustard-induced DNA damage and toxicity in skin keratinocytes
Influence of oxidative stress, diaphragm fatigue, and
Chronic dietary exposure of zebrafish to PAH mixtures results in carcinogenic but not genotoxic effects
Catalytic antioxidant AEOL 10150 treatment ameliorates sulfur mustard analog 2-chloroethyl ethyl sulfide-associated cutaneous toxic effects
Effects of seven chemicals on DNA damage in the rat urinary bladder: A comet assay study
Mitogen-activated protein kinase signal transduction and DNA repair network are involved in aluminum-induced DNA damage and adaptive response in root …
Sodium arsenite induced changes in survival, growth, metamorphosis and genotoxicity in the Indian cricket frog (< i> Rana limnocharis</i>)
Hypoxia Provokes Base Excision Repair Changes and a Repair-Deficient, Mutator Phenotype in Colorectal Cancer Cells
Monitoring regulation of DNA repair activities of cultured cells in-gel using the comet assay
Effects of Biovar I and Biovar II of< i> Ureaplasma urealyticum</i> on Sperm Parameters, Lipid Peroxidation, and Deoxyribonucleic Acid Damage in Male Infertility
MALDI-TOF fingerprinting of seminal plasma lipids in the study of human male infertility
Genotoxicity analysis of cerium oxide micro and nanoparticles in Wistar rats after 28 days of repeated oral administration
Sodium Fluoride Promotes Apoptosis by Generation of Reactive Oxygen Species in Human Lymphocytes
Genotoxicity assessment of cerium oxide nanoparticles in female Wistar rats after acute oral exposure
The differential localization of a methyl group confers to two triterpenes present in the olives a different anti-breast cancer activity.

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