Breast secretions can be classified into two types according to their major protein components. Type I fluids contain Zn-alpha 2-glycoprotein, apolipoprotein D, and gross cystic disease fluid protein-15, while Type II fluids are characterized by the presence of lactoferrin, lysozyme, and alpha-lactalbumin. In this study, the polypeptide composition of breast secretions from 719 nonlactating women was evaluated by using polyacrylamide gel electrophoresis. The required amount for the analysis (1 microliter) was obtained from 50% of control women and from 75% of women with mammary disease. There were more secretors in premenopausal than in postmenopausal women, as well as in parous than in nulliparous women. Evaluation of factors affecting protein composition of breast secretions revealed that Type II fluids were found in the majority of women who had given birth in the last four years and in a high proportion of oral contraceptive users. After excluding both of these groups, Type II fluids were detected in 47% of patients with breast cancer, but only in 8% of control women and in 16% of women with benign breast diseases. Taken together, these results suggest that protein analysis of breast secretions could be an useful tool for the study of breast pathologies.