Freehand three-dimensional ultrasound (3DUS) was used to investigate longitudinal and biaxial transverse deformation and rotation of the free Achilles tendon in vivo during a voluntary submaximal isometric muscle contraction. Participants (n = 8) were scanned at rest and during a 70% maximal voluntary isometric contraction (MVIC) of the plantarflexors. Ultrasound images were manually digitized to render a 3D reconstruction of the free Achilles tendon for the computation of tendon length, volume, cross-sectional area (CSA), mediolateral diameter (MLD), anteroposterior diameter (APD), and transverse rotation. Tendon longitudinal and transverse (CSA, APD, and MLD) deformation and strain at 70% MVIC were calculated relative to the resting condition. There was a significant main effect of contraction on tendon length and mean CSA, MLD, and APD (P < 0.05), but no effect on tendon volume (P = 0.70). Group mean transverse strains for CSA, MLD, and APD averaged over the length of the tendon were -5.5%, -8.7% and 8.7%, respectively. Peak CSA, MLD, and APD transverse strains all occurred between 40% and 60% of tendon length. Transverse rotation of the free tendon was negligible at rest but increased under load, becoming externally rotated relative to the calcaneal insertion. The relationship between longitudinal and transverse strains of the free Achilles tendon during muscle-induced elongation may be indicative of interfascicle reorganization. The finding that transverse rotation and strain peaked in midportion of the free Achilles tendon may have important implications for tendon injury mechanisms and estimation of tendon stress in vivo.