Comet assay is a quick and versatile technique for assessing DNA damage in individual cells. It allows the detection of DNA single- and double-strand breaks, as well as the presence of alkali-labile sites and cross-links. Here we describe protocols for the Single-Cell Gel Electrophoresis (Comet assay) in its alkaline (pH > 13), mild alkaline (pH = 12.1), and neutral (pH = 8) versions when applied in aquatic animals exposed to nanomaterials.