The zebrafish (Danio rerio) is a modelorganismwidely used in several research fields due to its characteristics and numerous advantages, such as optical embryo transparency, fully sequenced genome, orthologous genes to humans, smallsize, high reproductive rate, easy gene editingand relatively lowcosts. Thus, a number of protocols have been developed that allow the use of this vertebratemodel for toxic effect evaluation at various biological levels, including genotoxicity, using the comet assay technique.The comet assay or single-cell gel electrophoresis is a popular and sensitive method to study DNA damage in cells, which is described in this chapter. Briefly,cells suspended in agarose on a microscope slide are lysed, denatured, electrophoresed, neutralized, and stained to study the migration of DNA strand breaks. As a result, cells with increased DNA damage present a high fluorescence intensity and an increase ofcomet taillength. For the visual score, comets are classified according to the head integrity, tail intensity, and tail length into five classes, namely, class 0 until class 4 (comets with high damage and with almost all the DNA in the tail). These data are used to calculate the Genetic Damage Index (GDI) expressed as Arbitrary Units (AU).