11th INTERNATIONAL CONGRESS OF THE TURKISH SOCIETY OF TOXICOLOGY, Antalya, Türkiye, 02 Kasım 2022, ss.121-122
The comet assay providing the estimation of both DNA damage and repair is a fast, sensitive, and
affordable technique. For long years, DNA damage and repair in individual cells and tissues have
been evaluated by using the comet assay. Also, the comet assay is the standard method used in
ecogenotoxicology, molecular epidemiology, human and environmental biomonitoring. Single/
double DNA strand breaks, alkali labile sites, base-pair damages, and apoptotic nuclei can be
determined by using this method.
Positive controls are chemicals known to induce DNA damage by the way of direct or indirect, and
also they cause single/double DNA strand breaks identified by the comet assay. The examples of
positive controls used in the comet assay are hydrogen peroxide (H2O2), methyl methane sulfonate
(MMS), ethyl methane sulfonate (EMS), etoposide, and ethyl nitroso urea (ENU). The choice of a
suitable positive control is an essential part of genotoxicity tests. However, it has been stated that
the results obtained from different positive controls at discrete concentrations and time intervals
varied. Accordingly, we aimed to compare H2O2, and etoposide, at different concentrations for 30
minutes, and 1h, respectively.
In our study, the alkali comet assay (pH>13) was performed to compare the results obtained from
two different positive controls by using 3T3 cell lines. H2O2 was tested at 1, 5, 10, 25, 50, 75, 100
and 200 μM for 1h. For 30 minutes, up to 75 μM H2O2 concentrations were tested. As for etoposide,
six different concentrations (0,05, 0,1, 0,5, 1, 5, 10 μM) were used for 30 minutes and 1h.
According to the comet assay results, the exposure of 25 and 50 μM H2O2 for 30 minutes
significantly induced DNA damage more than for 60 minutes. DNA strand breaks resulted from
the exposure of 1, 5, and 10 μM etoposide concentrations for 30 minutes were higher than for 60
minutes (p<0,05, one-way ANOVA).