SUMMARY The goal of this proposal is to develop a quantitative framework to address sex as a biological variable in musculoskeletal modeling and simulation research. This new framework will allow the field to understand how sex differences in musculoskeletal structure influence movement biomechanics, musculoskeletal injury, and neuromuscular disease. There are many known differences in bone anatomy, joint mechanics, muscle architecture, and movement function between males and females. Likewise, it is well documented that there are significant sex differences in susceptibility to musculoskeletal injury and neuromuscular disease. However, while the use of computer simulations of movement to study how musculoskeletal structure influences neuromuscular injury and disease has increased dramatically due to advances in numerical algorithms and computational power, the models that are used are based on musculoskeletal data that (1) are derived from a male-only population, and/or (2) combine measures from males and females in a way that averages out any potential sex differences. These profound limitations leave the field without any tools to examine how the known sex differences in musculoskeletal structures may influence biomechanics, injury, and disease. This project has three key aims that will resolve these profound limitations. The first aim will develop a comprehensive digital database of lower limb muscles, joints, and bones across female and male populations of varying body sizes. This aim will be achieved through using high throughput image segmentation analysis of magnetic resonance images collected of 50 male and 50 female subjects. The second aim will incorporate the measurements from the first aim into a computational framework that enables for accurate sex-specific scaling of lower limb models, including the ability to capture the measured variability in the form of uncertainty analysis. The third aim will use the models in the second aim to develop a model-based analysis method to generate novel insights into sex differences in lower limb biomechanics. The analysis method will be applied to examine sex differences in muscle forces during walking and landing. Taken together, these aims will not only address critical questions related to differences in musculoskeletal structure and function between males and females, but also provide a rigorous, detailed, sex-specific digital database of data and models that will be provided open-access for the entire scientific community to use. This posted resource will empower the field with a set of tools to rigorously examine sex as a biological variable in musculoskeletal modeling research.