We investigated concentrations of prostate-specific antigen (PSA) in mid-stream urine of 213 patients. Among them were 34 females. Diagnoses of the male patients were 42 benign prostatic hypertrophy (BPH), 21 localized prostate cancer prior to radical prostatectomy (RP), 15 post-RP without distant or local recurrence, 5 post-RP with local recurrence and 82 with other urological diseases. PSA levels were determined by the Hybritech Tandem E method. Female urine samples were positive in 38% of the cases. This and the finding of PSA in urine of men after RP is most likely due to extraprostatic production by periurethral glands. Urinary PSA levels do not differ between patients with BPH, organ-confined prostate cancer and other diagnoses. In some cases, however, urine PSA levels can be elevated in men with local tumor recurrence after RP when serum levels are still undetectably low. This indicates that the determination of urinary PSA concentration might be useful in the follow-up of patients after RP.