To evaluate whether deficient DNA repair contributes to elevated DNA damage and breast carcinogenesis, we used the comet assay (single-cell alkaline gel electrophoresis) to measure the levels of DNA damage in peripheral lymphocytes from 70 breast cancer cases and 70 controls. DNA damage, measured as the comet tail moment, was not influenced by age, family history (FH), age at menarche, age at first birth or parity. The results showed that cancer cases had significantly higher DNA damage compared with controls; the comet tail moments (mean +/- SD) for cases and controls were: 10.78 +/- 3.63 and 6.86 +/- 2.76 (P < 0.001) for DNA damage at baseline (DB), 21.24 +/- 4.88 and 14.97 +/- 4.18 (P < 0.001) for DNA damage after exposure to 6 Gy of ionizing radiation (DIR), and 14.76 +/- 5.35 and 9.75 +/- 3.35 (P < 0.001) for DNA damage remaining after 10 min repair following exposure to 6 Gy of IR (DRP), respectively. Body mass index (BMI) affected DNA damage differently for cases and controls. Damage decreased with increasing BMI for controls, while damage increased with increasing BMI for cases. Above-median DNA damage was significantly associated with breast cancer risk; the age-adjusted odds ratio (OR) = 13.44 [95% confidence interval (CI) = 5.97-30.24] for DB, 13.65 (6.07-30.71) for DIR and 6.54 (3.11-13.79) for DRP, respectively. This association was stronger in women with above-median BMI. Our results, although based on a relatively small group of subjects, indicate that elevated DNA damage is significantly associated with breast cancer risk and warrant larger studies to further define the molecular mechanisms of DNA damage/repair in breast cancer susceptibility.