AbstractNew treatments for bone diseases require testing in animal models before clinical translation, and the mouse tibia is among the most common models. In vivo micro-Computed Tomography (microCT)-based micro-Finite Element (microFE) models can be used for predicting the bone strength non-invasively, after proper validation against experimental data. Different modelling techniques can be used to estimate the bone properties, and the accuracy associated with each is unclear. The aim of this study was to evaluate the ability of different microCT-based microFE models to predict the mechanical properties of the mouse tibia under compressive load. Twenty tibiae were microCT scanned at 10.4 µm voxel size and subsequently compressed at 0.03 mm/s until failure. Stiffness and failure load were measured from the load–displacement curves. Different microFE models were generated from each microCT image, with hexahedral or tetrahedral mesh, and homogeneous or heterogeneous material properties. Prediction accuracy was comparable among models. The best correlations between experimental and predicted mechanical properties, as well as lower errors, were obtained for hexahedral models with homogeneous material properties. Experimental stiffness and predicted stiffness were reasonably well correlated (R² = 0.53–0.65, average error of 13–17%). A lower correlation was found for failure load (R² = 0.21–0.48, average error of 9–15%). Experimental and predicted mechanical properties normalized by the total bone mass were strongly correlated (R² = 0.75–0.80 for stiffness, R² = 0.55–0.81 for failure load). In conclusion, hexahedral models with homogeneous material properties based on in vivo microCT images were shown to best predict the mechanical properties of the mouse tibia.