Background: According to PubMed, at least 3000 articles are published in the field of “blood vessels in muscle” every year. The skeletal muscles of the rodent hind limb are the most frequently used experimental model. However, most of the studies are based on a single muscle and and usually on solitary cross-sections. Taking into account that capillarization differs dramatically between different muscles and even within the same muscle, we decided to introduce a complex 3D assay of structure of the vasculature in the murine hind limb. Methods & Results: Although imaging and quantification of bones by µCT is well developed and established, visualization of striated muscle capillaries is a challenging issue, mainly due to lack of satisfactory contrast agents. Therefore, to visualize the 3D structure of the vascular network with a resolution up to the level of capillaries, vascular casting with PU4ii® polymer (vasQtec, Zurich) has been performed. To enhance the contrast, after maceration the casts have been immersed in osmium tetraoxide and freeze-dried thereafter and investigated with a µCT (Skyscan 1172, voxel size below 1µm). Additionally, special a histological approach has been employed. Since cutting of mineralized bone with an ordinary microtome is not feasible, the entire hind limb was decalcified in EDTA for 4 days. Subsequently, the limb was embedded in paraffin and series of 5µm-thick sections of the entire hind limb obtained. After deparaffinization and trypsinization of the tissue sections, the capillaries were stained with BS-1 lectin. This method allows comparative evaluation of the capillary-to-fibre ratio, capillary density, fibre type i) of all 11 muscles on a single cross-section; ii) at different levels in the same muscle, i.e. proximal to distal. Based on this method, we were able to show alterations among the above-mentioned parameters depending on the level examined. Implications: We present a comparative method to evaluate the vasculature of all skeletal muscles on a cross-section of the whole hind limb simultaneously and at different levels. This method allows a time-efficient, 3D and comparative evaluation of the hind limb vascularization.