The onset of anaerobic (lactate) metabolism during incremental exercise, which may be a result of an imbalance between tissue oxygen supply and demand, has been associated with the gas exchange ventilatory threshold (VT). This study was designed to examine whether near infrared spectroscopy (NIRS) could be used to detect the VT in healthy subjects. Twenty-one men and 19 women completed incremental cycle ergometry during which NIRS measurements were obtained from the right vastus lateralis and gas exchange measurements were monitored simultaneously using a metabolic cart. The VT was identified from the metabolic data by the V-slope method and from NIRS data as the intensity at which tissue absorbency crossed the resting baseline value observed immediately prior to the initiation of exercise. Pearson correlations for the relative oxygen uptake and power output observed for the two methods of detecting VT were 0.90 and 0.88, respectively, in men and 0.89 and 0.86, respectively, in women (P < 0.01). No significant differences were observed between the two methods of detecting VT for any of the physiological responses (P > 0.05). No significant (P > 0.05) gender differences were observed in muscle oxygenation values at the VT, 32% in men and 38% in women. These results validate the use of NIRS as an alternate noninvasive method for detecting VT during cycle exercise in healthy subjects.