The Comet Assay (Single-Cell Gel Test)

Abstract

The comet assay or single-cell gel (SCG) test is becoming established as a useful technique for studying DNA damage and repair. In this microgel electrophoresis technique, a small number of cells suspended in a thin agarose gel on a microscope slide is lysed, electrophoresed, and stained with a fluorescent DNA binding dye. Cells with increased DNA damage display increased migration of chromosomal DNA from the nucleus toward the anode, which resembles the shape of a comet (Fig. 1). In its alkaline version, which is mainly used, DNA strand breaks and alkali-labile sites become apparent, and the amount of DNA migration indicates the amount of DNA damage in the cell. The comet assay combines the simplicity of biochemical techniques for detecting DNA single-strand breaks and/or alkali-labile sites with the single-cell approach typical of cytogenetic assays. The advantages of the SCG test include its simple and rapid performance, its sensitivity for detecting DNA damage, the analysis of data at the level of the individual cell, the use of extremely small samples, and its applicability to virtually any eukaryote cell population. Apart from image analysis, which greatly facilitates and enhances the possibilities of comet measurements, the cost of performing the assay is extremely low. The comet assay has already been used in many studies to assess DNA damage and repair induced by various agents in a variety of cells in vitro and in vivo (for a review, see 1,2).