Publication date
2013Keyword
Comet assaySingle-cell electrophoresis
DNA damage
Double-strand breaks
Single-strand breaks
Alkali-labile sites
Peer-Reviewed
YesOpen Access status
closedAccess
Metadata
Show full item recordAbstract
The comet assay or single-cell gel electrophoresis assay is a relatively simple and sensitive technique for quantitatively measuring DNA damage and repair at the single-cell level in all types of tissue where a single-cell suspension can be obtained. Isolated cells are mixed with agarose, positioned on a glass slide, and then lysed in a high-salt solution which removes all cell contents except the nuclear matrix and DNA, which is finally subjected to electrophoresis. Damaged DNA is electrophoresed from the nuclear matrix into the agarose gel, resembling the appearance of a comet, while undamaged DNA remains largely within the proximity of the nuclear matrix. By choosing different pH conditions for electrophoresis, different damage types and levels of sensitivity are produced: a neutral (pH 8–9) electrophoresis mainly detects DNA double-strand breaks, while alkaline (pH ≥ 13) conditions detect double- and single-strand breaks as well as alkali-labile sites. This protocol describes a standard comet assay study for the analysis of DNA damage and outlines important variations of this protocol.Version
No full-text in the repositoryCitation
Anderson D and Laubenthal J (2013) Analysis of DNA damage via single-cell electrophoresis. In: Makovets S (Ed) DNA Electrophoresis. (Methods in Molecular Biology) New York: Humana Press: 209-218.Link to Version of Record
https://doi.org/10.1007/978-1-62703-565-1_14Type
Book chapterae974a485f413a2113503eed53cd6c53
https://doi.org/10.1007/978-1-62703-565-1_14