We examined the use of antibiotics among 2728 women with a first diagnosis of breast cancer during 1994-2003, and 27 280 population controls in North Jutland County, Denmark, based on hospital discharge diagnoses, prescription use from 1989 to 2002, and population registry data. We found no increased relative risk of breast cancer associated with use compared with nonuse. The odds ratio for breast cancer associated with more than 10 prescriptions for antibiotics was 1.00 (95% CI 0.86 -1.15). Relative risks were similar for different classes of antibiotics. A subanalysis based on cases and controls younger than 70 years of age, with data on first birth and number of children, showed similar risk estimates even after adjustment for age at first birth and parity. In our study, use of antibiotics was not associated with an increased risk of breast cancer.