1 This in vitro study was undertaken as a preliminary approach before assessing whether the alkaline elution assay can be applied to peripheral blood lymphocytes (PBL) for the monitoring of humans exposed to genotoxic agents such as polycyclic aromatic hydrocarbons (PAH). We have compared in vitro, with the aid of the alkaline elution assay, the formation and the repair of DNA single- strand breaks (ssb) induced by different genotoxic agents [gamma-irradiation, ethyl methanesulfonate (EMS), benzo<a>pyrene diol epoxide (BPDE)] on quiescent and PHA-stimulated human lymphocytes and on a fibroblast cell line. 2 Gamma-irradiation (4 Gy) induced an equivalent amount of DNA ssb in the three cell types. On the other hand, after treatment with EMS (10mM) and BPDE (50μM), a higher production of DNA ssb was observed in replicat ing cells (PHA-stimulated lymphocytes and fibroblasts) when compared with quiescent lymphocytes. 3 After gamma-irradiation, all cell types repaired more than 65% of ssb within 1 h. After treatment with EMS, we noted a deficient DNA repair capacity in quiescent lym phocytes in comparison with replicating cells. In all cell types treated with BPDE, more breaks were observed after a 2 h repair period than immediately after treatment, demonstrating the involvement of a slow repair mecha nism after BPDE treatment. 4 Several conclusions can be drawn from this pilot study, (i) when assessing in vitro the induction and the repair of DNA lesions induced by chemicals, it seems rea sonable to test both non-replicating and replicating cells since their response may be different; (ii) in view of the relative persistence of DNA damage induced in vitro by BPDE in resting lymphocytes, chronic exposure to PAH could give rise to a certain accumulation of DNA damage in coke oven workers lymphocytes. Further studies will be necessary to determine whether these damages could be detected by alkaline elution.
A Silver Staining Method for Single-cell Gel Assay