A new method is proposed for tracking individual motor units (MUs) across multiple experimental sessions on different days. The technique is based on a novel decomposition approach for high-density surface electromyography and was tested with two experimental studies for reliability and sensitivity. Experiment I (reliability): ten participants performed isometric knee extensions at 10, 30, 50 and 70% of their maximum voluntary contraction (MVC) force in three sessions, each separated by 1 week. Experiment II (sensitivity): seven participants performed 2 weeks of endurance training (cycling) and were tested pre–post intervention during isometric knee extensions at 10 and 30% MVC. The reliability (Experiment I) and sensitivity (Experiment II) of the measured MU properties were compared for the MUs tracked across sessions, with respect to all MUs identified in each session. In Experiment I, on average 38.3% and 40.1% of the identified MUs could be tracked across two sessions (1 and 2 weeks apart), for the vastus medialis and vastus lateralis, respectively. Moreover, the properties of the tracked MUs were more reliable across sessions than those of the full set of identified MUs (intra-class correlation coefficients ranged between 0.63—0.99 and 0.39–0.95, respectively). In Experiment II, ∼40% of the MUs could be tracked before and after the training intervention and training-induced changes in MU conduction velocity had an effect size of 2.1 (tracked MUs) and 1.5 (group of all identified motor units). These results show the possibility of monitoring MU properties longitudinally to document the effect of interventions or the progression of neuromuscular disorders.