Objective To evaluate the safety and efficacy of laparoscopic partial nephrectomy via a combined "trans-retro-peritoneal three-step" approach. Methods The clinical data of 32 patients who received laparoscopic partial nephrectomy between Jul. 2015 and Nov. 2015 were retrospectively analyzed. Twenty cases were via the traditional transperitoneal approach and the other 12 cases via a novel combined "trans-retro-peritoneal three-step" approach. The 3 steps in this novel approach included: (1) Through transperitoneal approach, Gerota's fascia and perirenal fat were incised to expose the tumor; (2) The peritoneum and Gerota's fascia were incised along the Told's line, and the renal artery was then separated on the surface of the psoas muscle; and (3) The renal artery was clamped, the edge of the tumor was marked, and finally the tumor was resected and the kidney was reconstructed. The perioperative data of the two groups were compared. Results The surgeries were successfully performed in all the 32 patients. Compared to traditional transperitoneal approach, our novel approach significantly shortened the operation time (P=0.014). The other perioperative parameters, such as estimated blood loss, were comparable in the two groups. Conclusion This combined "trans-retro-peritoneal three-step" approach has the combined the advantage of traditional transperitoneal (enough room for manipulation) and retroperitoneal (convenience to separate the renal arteries) approaches, especially when the renal artery is abnormal. Further studies are needed to compare the present approach with the traditional surgeries.