To investigate the reliability of quantitative analysis of dynamic stretching muscle stiffness using shear wave elas-tography (SWE), and to evaluate the influence of stretched levels and region of interest (ROI) sizes on the repeatability of SWE measurements. SWE videos of the gastrocnemius medius were collected during ankle movement from plantar flexion (PF) 40° to dorsiflexion (DF) 30°. Shear wave imageswere collected of ankle angles at PF 25°, 0°, DF 15°, and DF 30°, representing the slack status, mildly stretched level, moderately stretched level, and maximal stretched level of the gastrocnemius medius, respectively. ROI circles with diameters of 2 mm, 5 mm, and 8 mm were applied to measure the shear modulus. Intra-observer, and inter-observer repeatability of the measurements were compared among different stretched levels and ROI sizes. Twenty-one healthy volunteers were enrolled. Muscle stiffness increased as the ankle DF increased. Intraclass correlation coefficients (ICCs) of intra-observer and inter-observer repeatability obtained for ROI sizes of 2 mm, 5 mm and 8 mm indicated good to excellent repeatability at all stretched levels. Shear wave elastography appeared to be a reliable tool to evaluate the dynamic stretching muscle stiffness with satisfactory repeatability at various stretched levels of gastrocnemius medius. Good to excellent repeatability was found using different ROI sizes.