The aim of the study was to critically review the clinical approach to distinguish the sensory and motor nerve fascicles of the peripheral nerve system and to explore potential novel techniques to meet the clinical needs. The principles and shortcomings of the currently used methods for identification of sensory and motor nerve fascicles, including nerve morphology, electrical stimulation, spectroscopy, enzymohistochemistry staining (acetylcholinesterase [AchE], carbonic anhydrase [CA] and choline acetyltransferase [ChAC] histochemistry staining methods), and immunochemical staining were systematically reviewed. The progress in diffusion tensor imaging, proteomic approaches, and quantum dots (QDs) assessment in clinical applications to identify sensory or motor fascicles has been discussed. Traditional methods such as physical and enzymohistochemical methods are not suitable for the precise differentiation of sensory and motor nerve fascicles. Immunohistochemical staining using AchE, CA, and ChAC is promising in differentiation of sensory and motor nerve fascicles. Diffusion tensor imaging can reflect morphological details of nerve fibers. Proteomics can reveal the dynamics of specific proteins discriminating sensory and motor fascicles. QDs, with their size-dependent optical properties, make them the ideal protein markers for identification of the sensory or motor nerves. Diffusion tensor imaging, proteomics and QDs-imaging will facilitate the clinical identification of motor and sensory nerve fascicles, help in improving surgical success rates and assist in postoperative functional recovery.