The anatomy of appendicular bones has been demonstrated to be informative for taxonomic, paleontological, and functional studies of bats. Its use in such studies, however, is limited by the small number of skeletons available for this taxonomic group in scientific collections. Here we describe a protocol for the extraction of the femur from fluid-preserved bats. This protocol was successfully tested on a large sample, including 58 species in 43 genera and 9 families (a total of 183 specimens), and is minimally invasive, requiring only two incisions in the leg at disarticulation points (knee and coxofemoral joints). This method provides material for appendicular anatomical research with minimal damage to external morphology.