To determine the utility of fecal corticosteroid concentration as a measure of chronic stress under laboratory and field conditions, we biochemically and physiologically validated a radioimmunoassay for corticosteroids in three rodent species, house mice (Mus musculus), deer mice (Peromyscus maniculatus), and red-back voles (Clethrionomys gapperi). The biochemical validations demonstrated that the assay accurately and precisely measured corticosteroid concentration in the feces. The physiological validation indicated that the assay was sensitive enough to detect the stress associated with (a) brief handling and bleeding of animals, (b) chronic caloric restriction, (c) exposure to a novel environment, and (d) exposure to a novel cold environment. Our results suggest that fecal measurements reflect stress levels experienced by these animals approximately 6-12 h before defecation. Therefore, given a judicious trapping and trap-monitoring protocol, this assay has considerable utility for measuring the stress levels at which animals actually exist in the field.