The aim of this study is to compare different tools for evaluating prostate-specific antigen (PSA) increase or decrease, such as PSA velocity and PSA slope. This study was conducted on 312 male patients evaluated with transrectal ultrasound-guided biopsy of prostate with six or more cores. Patients with at least three consecutive PSA measurements in at least 18 months entered the study. Prostate-specific antigen slope was estimated by the slope of the least-square regression line fit to PSA versus time in years; PSA velocity was calculated with 3 or more PSA arrays. Median age was 66 years (range 45-86). Overall 67 patients were affected by primary prostate cancer, 245 were controls without prostate cancer. Prostate-specific antigen slope and PSA velocity were significantly higher in patients with prostate cancer than in controls. At the ROC analysis, PSA slope evidenced better results than PSA velocity (area under the curve (AUC) 0.743 for PSA slope; AUC 0.663 for PSA velocity; P=0.037). At PSA slope (calculated with the least-square fit) equal to zero, the sensitivity resulted as being 94% with a specificity of 38.8%. In conclusion prostate-specific antigen slope calculated with three or more PSA assays permits longitudinal evaluation of PSA for prostate diagnosis. Prostate-specific antigen slope improves both sensitivity and specificity in prostate cancer diagnosis, compared with PSA velocity.