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Many studies have revealed facts about the anatomy of the medial sural artery perforator. The exact locations or areas where the perforators pierce through the fascia, however, have not been clearly identified.Based on cadaver dissections performed on 40 legs from 20 cadavers, the authors identified the location of the first and second perforators of the medial sural artery. The passages of the medial sural artery perforators were also confirmed by means of duplex-Doppler scanning, three-dimensional computed tomography, and angiography.The authors found that the main perforators of the medial sural artery were located on a line drawn from the midpoint of the popliteal crease to the midpoint of the medial malleolus. The hip joint was flexed in supine position and the knee joint was flexed into a 90-degree angle, thus making it possible to observe the first perforator, which was approximately 8 cm from the midpoint of the popliteal crease, and the second perforator, which was approximately 15 cm from the popliteal crease. A series of 21 clinical cases with soft-tissue defects were successfully treated using the medial sural artery perforator flap, including 18 free flaps and three pedicled flaps. There were seven cases of hand reconstruction and 14 cases of lower extremity reconstruction.Until now, it has been a general belief that the exact location of a perforator cannot be predicted. The authors, however, have found that the first perforator was located almost 8 cm from the midpoint of the popliteal crease within the distal half circle drawn with a radius of 2 cm. Because this study made it possible to prepare a diagram of the exact location of the medial sural perforators, it is expected that more professionals will use this method for hand and facial reconstructions in the future.