The comet assay, also known as single-cell gel electrophoresis (SCGE), is a widely used method for detecting DNA strand breaks in eukaryotic cells. The assay involves embedding cells in agarose, lysing them with detergents and high salt, and then subjecting the DNA to electrophoresis at high pH. This process results in structures resembling comets, with the tail representing the DNA that has relaxed from supercoiling and the head containing the more intact DNA. The intensity of the comet tail relative to the head reflects the number of DNA breaks. The assay has several variants, including alkaline and neutral SCGE, each with its own advantages and sensitivities. Alkaline SCGE is commonly used due to its ability to detect a wider range of damage, while neutral SCGE is more sensitive to single-strand breaks. The introduction of lesion-specific endonucleases further enhances the assay's specificity, allowing the detection of specific types of DNA damage such as UV-induced pyrimidine dimers, oxidized bases, and alkylation damage. The comet assay is widely applied in genotoxicity testing, human biomonitoring, molecular epidemiology, and ecological monitoring. It can be used to assess the genotoxicity of chemicals, monitor environmental contamination, and investigate the effects of dietary factors on DNA damage. The assay is also useful in diagnosing genetic disorders and assessing background levels of DNA damage. Quantitation of the comet assay involves visual scoring or image analysis, both of which provide valuable information about the extent of DNA damage. The assay can be standardized and quantified to ensure reliability, and it has been shown to be useful in both human population studies and animal experiments. However, it is important to consider factors such as cell viability, storage conditions, and statistical analysis to ensure accurate results. Overall, the comet assay is a powerful tool for studying DNA damage and repair, offering insights into the mechanisms of genotoxicity and the variation in DNA repair capacity among individuals.The comet assay, also known as single-cell gel electrophoresis (SCGE), is a widely used method for detecting DNA strand breaks in eukaryotic cells. The assay involves embedding cells in agarose, lysing them with detergents and high salt, and then subjecting the DNA to electrophoresis at high pH. This process results in structures resembling comets, with the tail representing the DNA that has relaxed from supercoiling and the head containing the more intact DNA. The intensity of the comet tail relative to the head reflects the number of DNA breaks. The assay has several variants, including alkaline and neutral SCGE, each with its own advantages and sensitivities. Alkaline SCGE is commonly used due to its ability to detect a wider range of damage, while neutral SCGE is more sensitive to single-strand breaks. The introduction of lesion-specific endonucleases further enhances the assay's specificity, allowing the detection of specific types of DNA damage such as UV-induced pyrimidine dimers, oxidized bases, and alkylation damage. The comet assay is widely applied in genotoxicity testing, human biomonitoring, molecular epidemiology, and ecological monitoring. It can be used to assess the genotoxicity of chemicals, monitor environmental contamination, and investigate the effects of dietary factors on DNA damage. The assay is also useful in diagnosing genetic disorders and assessing background levels of DNA damage. Quantitation of the comet assay involves visual scoring or image analysis, both of which provide valuable information about the extent of DNA damage. The assay can be standardized and quantified to ensure reliability, and it has been shown to be useful in both human population studies and animal experiments. However, it is important to consider factors such as cell viability, storage conditions, and statistical analysis to ensure accurate results. Overall, the comet assay is a powerful tool for studying DNA damage and repair, offering insights into the mechanisms of genotoxicity and the variation in DNA repair capacity among individuals.