Complete removal of the tumor by surgery offers the best chance for cancer cure; however, many prostate cancer patients who have negative surgical margins at radical prostatectomy will still experience local and distant tumor recurrence. In other organs, the closest distance between tumor and resection margin has prognostic significance. This has not been adequately studied in prostatectomy specimens. We undertook a prospective study of 278 consecutive margin-negative whole-mount prostatectomy cases. The anatomic location and closest distance between tumor and resection margin, measured with an ocular micrometer, were analyzed. All the slides were reviewed by a single pathologist, and data were collected prospectively. The closest distance between tumor and resection margin ranged from 0.02 to 5.0 mm (mean, 0.7 mm; median, 0.5 mm) and correlated with patient age (P = 0.03), prostate weight (P = 0.002), Gleason score (P = 0.001), pathologic stage (P = 0.01), tumor volume (P < 0.001), and perineural invasion (P < 0.001). The closest distance between tumor and resection margin was not a significant predictor of PSA recurrence in univariate or multivariate logistic regression; and we do not, therefore, advocate reporting the closest distance between tumor and resection margin as a standard part of the surgical pathology report on prostatectomy specimens.