Breast cancer is believed to develop as multiple genetic abnormalities accumulate, each conferring some growth advantage, but the timing and nature of the earliest steps in this progression are not yet elucidated. Proliferative breast lesions, associated with an increased risk of breast cancer although considered benign, recently were shown to contain clonal genetic abnormalities. Therefore, we hypothesized that clonal genetic abnormalities might be detectable before any phenotypic abnormalities are evident, ie, in histologically normal breast tissue. We examined DNA extracted from 95 normal-appearing breast ducts or terminal ductal-lobular units from 20 individuals at varying degrees of risk (those undergoing reduction mammoplasties, those with atypical hyperplastic proliferative lesions, and those already diagnosed with breast cancer). Using nine microsatellite markers, we sought evidence of genetic instability or of allelic imbalance (most likely representing loss of heterozygosity). We found genetically abnormal clones in 21/95 (22%) seemingly normal samples from 10/20 (50%) women from all three risk groups. In women under age 50, trends toward increased rates of abnormalities were noted with increased cancer risk. The abnormalities identified were more likely to be at sites of known or postulated tumor suppressor genes rather than at random or neutral loci. Our data indicate that genetic abnormalities potentially critical to breast tumorigenesis accumulate before pathological detection even of high-risk lesions and are detectable in tissue that is not only histologically benign but also completely normal.