We investigated whether the free-to-total prostate-specific antigen (PSA) ration (f-PSA/t-PSA ratio; i.e. percentage of free PSA) represents a better discriminator for the detection of cancer of the prostate (CaP). In a retrospective analysis, the percentage of free PSA was determined in the sera of 35 patients with histologically proven benign prostatic hyperplasia (BPH) and 35 patients with clinically localized CaP. Patients with urolithiasis (n = 33) served as a control group. Serum levels of free PSA and total PSA were determined employing a chemiluminescent enzyme immunoassay. Patients with CaP demonstrated a lower percentage of free PSA (median: 8.7) than patients with BPH (median: 20.0; P < 0.001). Determination of the percentage of free PSA enhances the differentiation between BPH and CaP and may reduce the number of unnecessary biopsies in patients with an elevated PSA. Confirmation of our preliminary results is required.