Near-infrared spectroscopy (NIRS) is a non-invasive optical imaging technique, which is increasingly used to measure hemodynamic responses in the motor cortex. The location at which the NIRS optodes are placed on the skull is a major factor in measuring the hemodynamic responses optimally. In this study, the validity of using transcranial magnetic stimulation (TMS) in combination with a 3D motion analysis system to relocate the TMS derived position was tested. In addition, the main goal was to quantify the advantage of using TMS to locate the optimal position in relation to the most commonly used EEG C3 position. Markers were placed on the TMS coil and on the head of the subject. In eleven subjects, a TMS measurement was performed to determine the individual motor-evoked potential center-of-gravity (MEP-CoG). This procedure was repeated in nine subjects to test the validity. Subsequently, hemodynamic responses were measured at the MEP-CoG position and at the C3 position during a thumb abduction and adduction task. On average, the MEP-CoG location was located 19.2. mm away from the C3 position. The reproducibility study on the MEP-CoG relocation procedure revealed no systematic relocations. No differences in early and delayed hemodynamic responses were found between the C3 and MEP-CoG position. These results indicate that using TMS for NIRS optodes positioning on the motor cortex does not result in higher hemodynamic response amplitudes. This could be explained if NIRS and TMS assess slightly different functions. © 2011 Elsevier B.V.