The spacing between the median nerve and transverse carpal ligament (TCL) within the carpal tunnel can potentially affect the nerve morphology. This study aimed to quantify the spatial relationship between the median nerve and transverse carpal ligament in asymptomatic hands. Twelve subjects were recruited to image the carpal tunnel using robot-assisted ultrasound. The median nerve and TCL were segmented from each image and three-dimensionally reconstructed using kinematic information from the robot. The TCL-median nerve distance, nerve cross-sectional area, circularity, and position were measured along the entirety of the nerve length within the carpal tunnel. Results were averaged at every 5% of nerve length. At the nerve length percentages of 0% (distal), 25%, 50%, 75%, and 100% (proximal), the TCL-median nerve distance (±SD) was 0.7 ± 0.4, 0.7 ± 0.2, 0.5 ± 0.2, 0.5 ± 0.2, and 0.6 ± 0.3 mm, respectively. The corresponding nerve cross-sectional area was 9.4 ± 1.9, 10.6 ± 2.6, 11.2 ± 2.1, 11.2 ± 1.7, and 9.7 ± 1.9 mm2. A one-way analysis of variance showed no significant differences between the respective percentages of nerve length for TCL-median nerve distance (p = 0.219) and cross-sectional area (p = 0.869). Significant (p < 0.0001) but weak correlations were observed between the TCL-median nerve distance with cross-sectional area (r = −0.247) and circularity (r = −0.244). This study shows that the healthy median nerve morphology is consistent along the continuous nerve length within the carpal tunnel, supporting the use of 2D imaging in the evaluation of the healthy nerve.