Following the widespread use of radioguided surgery (RGS) in melanoma and breast cancer, we applied this new surgical strategy to prostate cancer (PC). The aims of this study were 1) to evaluate the accuracy of RGS in the detection of prostatic sentinel lymph nodes (SLN), and 2) to verify if pelvic lymphadenectomy (LAD) is an accurate means to detect solitary micrometastases. We investigated 48 patients with PC confirmed by transrectal biopsy who underwent radical prostatectomy and bilateral LAD. A dose of 99mTc-labeled nanocolloid particles was injected into the prostate after needle positioning by ultrasonography. Serial imaging was obtained with a gamma camera, identifying 1) the first radioactive lymph node (sentinel lymph node, SLN); 2) other radioactive lymph nodes, and 3) non-active lymph nodes. Forty-three SLNs were identified in 48 patients. Twenty SLNs were located at unusual sites with respect to the extent of conventional LAD. Five SLNs were positive for micrometastases and two of these were located outside the usual LAD area. No micrometastases were found in any of the remaining lymph nodes (active and non-active). These preliminary results are in agreement with the few previous scientific contributions available on this topic and indicate that it is possible to reduce the extent and duration of surgery and necessary to reevaluate the conventional sites of lymphatic drainage.