BACKGROUND: Although emotion-specific autonomic responses based on the discrete theory of emotion have been widely studied, studies on the reliability of physiological responses to emotional stimuli are limited. In this study, we aimed to assess the reliability of physiological changes induced by the six basic emotions (happiness, sadness, anger, fear, disgust, and surprise) that were measured during 10 weekly repeated experiments. METHODS: Twelve college students participated, and in each experiment, physiological signals were collected before and while participants were watching emotion-provoking film clips. Additionally, the participants self-evaluated the emotions that they experienced during the film presentation at the end of each emotional stimulus. To avoid adaptation of participants to identical stimuli during repeated measurements, we used 10 different film clips for each emotion, and thus a total of 60 film clips over 10 weeks were used. Physiological features, such as skin conductance level (SCL), fingertip temperature (FT), heart rate (HR), and blood volume pulse (BVP), were extracted from the physiological signals. Two reliability indices, Cronbach's alpha and intraclass correlation coefficient, were calculated from the physiological features to assess internal consistency and interrater reliability, respectively. RESULTS: We found that SCL, HR, and BVP measured during the emotion-provoking phase over the 10 weekly sessions were more reliable than those assessed at baseline. Furthermore, SCL, HR, and BVP from the emotion-provoking phase exhibited excellent internal consistency and interrater reliability. CONCLUSIONS: Our findings suggest that these features can be used as reliable physiological indices in emotion studies. The results also support the significance of physiological signals as meaningful indicators for emotion recognition in HCI (human computer interface) area.