Neuromuscular disorders frequently complicate sepsis and other critical illnesses in patients. Mice are the major species used as a model for sepsis. Nerve conduction studies (NCS), the primary tool for noninvasive assessment of nerve and muscle function, is challenging to perform in small animals. A reliable method for noninvasive, repeated NCS testing has not been reported in mice. We developed and validated a method for the repeated measurement of mouse sciatic nerve conduction in normal and septic mice. Our sedated and awake NCS system enabled minimally invasive long-term repeated measurements. The mean compound muscle action potential (CMAP) amplitude and latency were 17.4 mV and 1.11 ms, respectively (n = 59). There was an excellent intertester reproducibility by linear regression in both normal (r = 0.95) and septic (r = 0.98) mice. We also showed significant, time-dependent isoflurane-induced CMAP suppression in all animals, which was further exacerbated in septic mice. This study provides a new tool for the assessment of peripheral nerve/muscle function in mouse neuromuscular disease models that require repeated, long-term, and minimally invasive monitoring.