The role of HER-2/ neu in male breast cancer is not well defined. The purpose of the current study was to measure the frequency of HER-2/ neu expression in primary male breast cancer, to demonstrate HER-2/ neu gene amplification in cases found to be positive for protein overexpression, and to correlate HER-2/ neu positivity with clinicopathological variables. Formalin-fixed, paraffin-embedded archival material from 99 primary male breast carcinomas was evaluated by immunohistochemistry (IHC) using the HercepTest (DAKO Corp., Hamburg, Germany). Scoring was performed according to established guidelines. All cases demonstrating HER-2/ neu staining by IHC (1+/2+ and 3+) were analyzed for HER-2/ neu gene amplification by fluorescence in situ hybridization (FISH) utilizing the PathVysion assay (Vysis Corp., Downers Grove, Illinois) to assess HER-2/ neu amplification status. The immunohistochemical staining of the HER-2/ neu protein revealed HER-2/ neu positivity in 15/99 (15.1%) cases, eight tumors showed 2+ and 7 tumors 3+ staining. HER-2/ neu gene amplification was observed in 11/99 cases (11,1%), and all of the 3+ and 4/8 from the 2+ cases were amplified. HER-2/ neu gene amplification/protein overexpression did not correlate with tumor state, histological grade or estrogen/progesterone receptor status nor the axillary lymph node status. This is the first comprehensive study of HER-2/ neu gene amplification by FISH analysis in primary male breast cancer. Compared to female primary breast cancer the percentage of HER-2/ neu positivity in our study was lower. Our data provide first evidence for HER-2/ neu gene amplification in male breast cancer. Further studies should be addressed on the potential application of the monoclonal rhuMAB HER-2/ neu antibody for treatment of HER-2/ neu positive male breast cancer.