The aim of this study was to compare the use of transcutaneous versus motor nerve stimulation in the evaluation of low-frequency fatigue. Nine female and eleven male subjects, all physically active performed a 30-min downhill run on a motorized treadmill. Knee extensor muscle (KE) contractile characteristics were measured prior to, immediately following (POST) and 30 min after the fatiguing exercise (POST30) using single twitches, and 0.5 s tetani at 20 Hz (P20) and 80 Hz (P80). The P20^.P80^-1 ratio was calculated. Electrical stimulations were randomly applied either maximally to the femoral nerve (NES) or using large surface electrodes (ES) at an intensity sufficient to evoke 50% of MVC during a 80 Hz-tetanus. Voluntary activation level (%VA) was also determined during isometric maximal voluntary contraction (MVC) using the twitch-interpolation technique. KE MVC and %VA decreased at all points in time post exercise (P<0.001). P20 and P80 displayed significant time x gender x stimulation method interactions (P<0.05 and P<0.001, respectively). Both stimulation methods detected significant torque reductions at POST and POST30. Overall, ES tended to detect a greater impairment at POST in male and a lesser one in female subjects at both POST and POST30. Interestingly, the P20^.P80^-1 ratio relative decrease did not differ between the two methods of stimulation. The low-to-high frequency ratio only demonstrated a significant time effect (P<0.001). It can be concluded that low-frequency fatigue due to eccentric exercise appears to be accurately assessable using ES.