This paper describes the use of a computational method based on an optical flow algorithm to detect regions of interest in near infrared (NIR) spectroscopy. The evaluation of such method is also presented. Visual inspection and cross correlation analysis of NIR cortical activation images were used to evaluate our method. The visual analysis exposed pain-related activations in the primary somatosensory cortex (S1) after stimulation which is consistent with similar studies, and the cross correlation results showed dominant channels on both cerebral hemispheres. Optical flow exhibited the nature of the dominant channel, the extent of the stimulation spatial distribution and the stimulation status. In addition, the directions of the optical flow vectors were linked to the stimulation perception of the participant. The two evaluation methods confirmed the success of our method in finding the region of interest in both hemispheres. The results of this real case study show that the computational method can successfully analyse and detect regions of interest and show temporal interactions between channels, and could be employed to investigate pain assessment in human subjects.