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Long‐term noninvasive sampling for endangered or elusive species is particularly difficult due to the challenge of collecting fecal samples before hormone metabolite desiccation, as well as the difficulty in collecting a large enough sample size from all individuals. Hair samples may provide an environmentally stable alternative that provides a long‐term assessment of stress and reproductive hormone profiles for captive, zoo, and wild mammals. Here, we extracted and analyzed both cortisol and testosterone in coyote (Canis latrans) hair for the first time. We collected samples from 5‐week old coyote pups (six female, six male) housed at the USDA‐NWRC Predator Research Facility in Millville, UT. Each individual pup was shaved in six different locations to assess variation in concentrations by body region. We found that pup hair cortisol (F_5,57.1 = 0.47, p = 0.80) and testosterone concentrations (F_5,60 = 1.03 …