Cytokinesis-block micronucleus cytome assay in lymphocytes

The cytokinesis-block micronucleus cytome (CBMN cyt) assay is a new and comprehensive technique for measuring DNA damage, cytostasis, and cytotoxicity in different tissue types, including lymphocytes. DNA damage events are scored specifically in once-divided binucleated cells. These events include; (a) micro-nuclei (MNi), a biomarker of chromosome breakage and/or whole chromosome loss; (b) nucleoplasmic bridges (NPBs), a biomarker of DNA misrepair and/or telomere end-fusions; and (c) nuclear buds (NBUDs), a biomarker of elimination of amplified DNA and/or DNA repair complexes. Cytostatic effects are measured via the proportion of mono-, bi-, and multinucleated cells and cytotoxicity via necrotic and/or apoptotic cell ratios. The assay has been applied to the biomonitoring of in vivo exposure to genotoxins, in vitro genotoxicity testing and in diverse research fields, such as nutrigenomics and pharmacogenomics. It has also been shown to be important as a predictor of normal tissue and tumor radiation sensitivity and cancer risk. This protocol also describes the current established methods for culturing lymphocytes, slide preparation, cellular and nuclear staining, scoring criteria, data recording, and analyses.