The purpose of this investigation was to determine whether there is gender difference in sleep comfort of healthy individuals at various temperatures. During winter, sleep quality was examined under different indoor temperatures (17, 20 and 23°C) using questionnaires and electroencephalogram (EEG). To explore the mechanism responsible for gender differences in comfortable sleeping temperatures, mean skin temperature, finger temperature and finger blood flow were measured. The results showed that females would prefer a higher ambient temperature during sleep than the men. The mean skin temperature for females was higher than that of males, whereas finger skin temperature and finger blood flow were significantly lower in females than in males. Furthermore, skin temperature and finger blood flow were more sensitive to ambient temperature with females than in males. The gender differences in preferred sleeping temperature could therefore be related to these physiological characteristics. Both subjective evaluations and EEG found better sleep quality in males under the same temperatures compared to females. Skin temperature changes over the course of the night also demonstrated longer periods of deep sleep in males compared to females.