In this study, we investigated the relationship between the activity rhythms of <italic>Camponotus japonicus</italic> worker ants and their interactions. Specifically, one or two workers collected from either inside or outside the nest in a breeding colony were placed in a measurement system under a constant dark condition, and their activity rhythms were measured for 14 days. We thereby examined the relationship between the activity rhythm in the system and the experimental conditions, which consisted of four different combinations of working locations during breeding (in/outside the nest) and single/double workers (one ant / two ants) in the measurement system, over a total of 96 samples. A large number of the sampled ants (about 90% of the total) showed circadian activity rhythms. The proportion of circadian activity rhythm was lower and the dispersion of the period was larger in the circadian activity rhythm observed in single workers collected from within the nest than in the other three experimental conditions. In all four experimental conditions, the amplitude of the circadian activity rhythm decayed on an approximate 5-day scale. These results provide quantitative evidence that the activity rhythm of ants is determined by the location of labor and individual interactions during breeding.