In vivo and in vitro cell populations exhibit heterogeneous responses to many genotoxic agents which can be partially explained by the relative position of the cells in the mitotic cycle. In this study, the possibility to simultaneously detect oxidative DNA damage and cell cycle position has been investigated in C3H10T1/2 cell line. Exponential and plateau phase cells were treated with hydrogen peroxide and DNA damage and repair were evaluated by the alkaline comet assay and the micronucleus test. Our results show a dose-related increase of DNA damage after H2O2 treatment in both culture conditions. The percentage of cells in the various phases of cell cycle determined by comet assay was compared with the data obtained by DNA content flow cytometric analysis, and a good agreement between the results of the two techniques was found. Untreated exponentially growing cells in G1 phase showed a lower tail moment than S and G2/M cells. The same cell cycle dependence was evidenced in cells treated with low doses of H2O2, while, at higher doses, all cells showed a similar high level of damage. DNA repair was assessed analyzing cells by comet assay 2 h after treatment. The results showed that DNA repair was almost complete for all H2O2 doses tested. These results confirm the sensitivity of the Comet Assay in assessing DNA damage and repair, and support its usefulness in evaluating cell cycle-related differential sensitivity to genotoxic agents.

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