A change in Y chromosome number is but one of the many cytogenetic abnormalities reported in human prostate tumors. However, reports in the literature have varied regarding the frequency of Y loss or gain, whether it is restricted to the cancerous tissue, and its relation to the biology of the disease. The most frequently used materials for analysis of Y enumeration have been metaphase spreads from short-term cell cultures of prostate tumor tissue and paraffin-embedded tissue sections. Analysis of Y chromosome number by using metaphase spreads on short-term cultures can be misleading owing to clonal cell selection during the establishment of these cultures. This may result in an incomplete representation of the loss/gain pattern in the tumor as a whole. Studies using paraffin-embedded tissue sections can be complicated by apparent chromosome loss due to nuclear truncation as a result of tumor sectioning. In an attempt to circumvent these problems, we have used touch preparations from human prostate tumors to search for Y chromosome loss. Fluorescence in situ hybridization analysis was conducted by using a whole chromosome Y paint, with an alpha-satellite chromosome 3 probe as a control, on tumor samples from 42 patients ages 40-75. The results demonstrated a gain of Y in a single prostate tumor sample, with no convincing evidence for loss of the entire Y chromosome in any of the other 41 samples examined. The results suggest that loss of the entire Y chromosome is an infrequent event in prostate cancer.