Although a splicing variant of prostate specific antigen (PSA-v) mRNA has been described previously, whether its protein (PSA-v or PSA-related protein 1, i.e., PSA-RP1) is actually expressed in human prostate cells remains elusive. We report that PSA-v protein is expressed in prostatic epithelia of both cancerous and benign tissues. Also, secreted PSA-v can be detected in the medium of a prostate cancer (PCa) cell line. Consistently, PSA-v mRNA is exclusively expressed in benign luminal epithelia and cancer cells of the prostate by in situ hybridization. Northern analysis of a cohort of 51 pairs of RNA samples from microdissected tissues showed that PSA-v mRNA levels remained constant in both benign and cancerous tissues, whereas PSA levels declined in cancerous areas. Our result suggests that it would be feasible to develop proper immunoassays for PSA-v to test whether PSA-v could have some clinical utility.