Purpose: Graduated compressive stockings (GCS) have been suggested to influence performance by reducing muscle oscillations and improving muscle function and efficiency. However, no study to date has analyzed the influence of GCS on muscle activity during running. The objective of the study was to analyze the influence of GCS on the perception of comfort and muscle activation of the main muscles of the lower leg during running. Method: Thirty-six participants ran on a treadmill with (GCS) or without (control) GCS. The running tests consisted of a 10-min warm-up followed by a 20-min intense run at 75% of the athlete's maximal aerobic speed. Surface electromyography of the tibialis anterior, peroneus longus, gastrocnemius lateralis (GL), and gastrocnemius medialis (GM) were recorded every 5 min during the run and analyzed using a non-linearly scaled wavelet analysis. Perception of comfort of the GCS was measured before and after the run. Results: The GCS were reported as comfortable garments and reduced GL activity at Minute 0 (p < 0.05, ?[superscript 2][subscript p] = 0.245) and Minute 5 (p < 0.05, ?[superscript 2][subscript p] = 0.326) and GM activity at Minute 0 (p < 0.05, ?[superscript 2][subscript p] = 0.233) compared with running without garments, but their effect was temporary and disappeared after 5 min of running. Conclusion: Even though GCS reduced gastrocnemius muscle activity during the initial minutes of running, it is hypothesized that the GCS could have lost their initial levels of compression after some minutes of exercise, thereby reducing their influence on muscle activation. However, this hypothesis needs to be further investigated.