The chromosomal aberration test is a standard assay used to detect genetic damage in eukaryotic cells, which can be caused by various chemical agents, environmental factors, or radiation. This protocol outlines the procedures for the treatment of cells with potential genotoxic agents and the subsequent collection of cells for chromosomal aberration analysis. Cells are exposed to the test substance under controlled conditions, and mitotic cells are harvested at specific time points to assess chromosomal structural changes. The protocol includes details on cell culture, treatment duration, metaphase arrest using colcemid, cell harvesting, slide preparation, and chromosomal analysis. This method provides a reliable approach for evaluating the genotoxic potential of substances by identifying structural chromosomal aberrations, including breaks, gaps, and translocations.