This review aims to comprehensively summarize the application of electrophysiological methods, specifically repetitive nerve stimulation (RNS) and single fiber electromyography (SFEMG), in the diagnosis of neuromuscular junction (NMJ) disorders, including myasthenia gravis, Lambert-Eaton syndrome, and sarcopenia in the elderly. Both RNS and SFEMG have demonstrated high sensitivity and specificity in detecting NMJ abnormalities. RNS aids in distinguishing presynaptic from postsynaptic lesions, while SFEMG provides direct evidence of NMJ function by assessing single motor unit action potentials. Key parameters in SFEMG, such as fiber density, jitter, and pulse blocking, are crucial for evaluating NMJ function. Increased fiber density and jitter value, along with pulse blocking, are often observed in patients with NMJ disorders. However, despite the extensive application of these techniques in various NMJrelated diseases, their role in aging, particularly in sarcopenic patients, remains underexplored, highlighting the need for future research.