Intramuscular injection of the calpain inhibitor leupeptin promotes peripheral nerve regeneration in primates (Badalamente et al., 1989), and direct positive effects of leupeptin on axon outgrowth were observed in vitro (Hausott et al., 2012). In this study, we applied leupeptin (2mg/ml) directly to collagen-filled nerve conduits in the rat sciatic nerve transection model. Analysis of myelinated axons and retrogradely labeled motoneurons as well as functional 'catwalk' video analysis did not reveal significant differences between vehicle controls and leupeptin treated animals. Therefore, leupeptin does not improve nerve regeneration via protease inhibition in regrowing axons or in surrounding Schwann cells following a single application to a peripheral nerve conduit suggesting indirect effects on motor endplate integrity if applied systemically.