The objective of this study was to determine if a transcranial magnetic stimulation (TMS) method of quantifying the degree to which the motor cortex drives the muscles during voluntary efforts can be reliably applied to the human knee extensors. Although the technique for estimating “cortical” voluntary activation (VA) is valid and reliable for elbow flexors and wrist extensors, evidence that it can be applied to muscles of the lower limb is necessary if twitch interpolation with TMS is to be widely used in research or clinical practice. Eight subjects completed two identical test sessions involving brief isometric knee extensions at forces ranging from rest to maximal voluntary contraction (MVC). Electromyographic (EMG) responses to TMS of the motor cortex and electrical stimulation of the femoral nerve were recorded from the rectus femoris (RF) and biceps femoris (BF) muscles, and knee extension twitch forces evoked by stimulation were measured. The amplitude of TMS‐evoked twitch forces decreased linearly between 25% and 100% MVC (r² > 0.9), and produced reliable estimations of resting twitch and VA (ICC_2,1 > 0.85). The reliability and size of cortical measures of VA were comparable to those derived from motor nerve stimulation when the resting twitches were estimated on the basis of as few as three TMS trials. Thus, TMS measures of VA may provide a reliable and valid tool in studies investigating central fatigue due to exercise and neurological deficits in neural drive in the lower limbs. © 2008 Wiley Periodicals, Inc. Muscle Nerve 2008