Thermal comfort in an office impacts physical health, stress, and productivity. Humidity affects thermal comfort; however, the underlying mechanism remains unclear. This study assessed the influence of humidity on body temperature, thermal comfort, stress, and their relationship in working individuals. Thirteen participants performed three sets of 20-min calculation tasks followed by a 10-min rest in 26 °C or 33 °C with relative humidity (RH) of 30 % or 60 %. Core body temperature (T), mean skin surface temperature (T), and electrocardiogram were continuously recorded. Subjective thermal sensations and comfort were assessed with visual analog scales. Stress level was estimated based on α-amylase activity and immunoglobulin A level in saliva and heart rate variability. Mean T and T elevated at 33 °C with 60 % RH, where warm sensation and thermal discomfort also increased. Heart rate variability reflecting parasympathetic nerve activity decreased. There was a negative linear relationship between weighted body temperature and thermal comfort. However, thermal discomfort was augmented at a given weighted body temperature at 60 % RH. Thus, under indoor working conditions, high humidity may augment thermal discomfort and become a stress factor. Increases in T and T are involved in the mechanism, alongside other factors.