The semitendinosus, one of the hamstring muscles, is characterized by a tendinous inscription separating the muscle into two distinct neuromuscular compartments. Additionally, its long distal tendon is prime autologous graft tissue for surgical reconstructions, such as anterior cruciate ligament surgery. However, the structure and function of semitendinosus compartments has not been well-documented in vivo, and how compartment morphology may potentially change after tendon harvesting remains unknown. In this thesis, the structure and function of the semitendinosus was examined using a variety of medical imaging techniques and musculoskeletal modelling.[...] Overall, studies conducted for this thesis indicated that assessments of the hamstrings, particularly the semitendinosus after anterior cruciate ligament reconstruction, have much room for improvement. Namely, ultrasonographic methods provide a more accessible manner for assessing hamstring function, more care needs to be taken when recording surface electromyograms from semitendinosus, functional assessments of individuals recovering from anterior cruciate ligament reconstruction must include isometric tests at 90° of knee flexion, and researchers need to incorporate more appropriate passive length-tension curves in musculoskeletal models. Moreover, the semitendinosus is comprised of two distinct neuromuscular compartments that differ in proximodistal length, but not radial size. This atypical structure for an extremity muscle should be accounted for in future investigations, as there may be crucial functional relevance for this split design. The cross-sectional nature of studies and the lack of measurement of the contractile component of semitendinosus compartments during both passive and active tasks precludes concrete conclusions on the structure, function, and adaptations to the two compartments in healthy and anterior cruciate ligament reconstructed legs. Nonetheless, the results obtained in this thesis have direct clinical benefit and generate hypotheses and ideas for future investigations to better uncover the origin and functional relevance of the unique design of the semitendinosus muscle.