Glucocorticoid (GC) hormones, provide basic information on endocrine activity as a function of interactions with their environment. When faced with short-term stressors, GCs elevate to mobilize glucose and coordinate activity of other metabolic processes as an adaptive response to the stressor. A multi-year project with the Missouri Department of Conservation aims to evaluate health and population responses of bats by comparing them on sites specifically managed for the endangered Indiana Bat, to those on reference sites not so managed. Our health goal is to obtain longer-term measures of cortisol, the main GC in microbats, from hair samples, and compare young-of-the-year bats across sites to give insight to potential differences in stress due to management practices. To refine our methods we focused on samples from Big Brown Bat, Eptesicus fuscus (EPFU), a relatively well-studied North American species. To start, we first pooled male EPFU hair samples from adult males, prepared the hair for hormone extraction by performing a series of ethanol washes, evaporating, and homogenizing. We then extracted known weights of hair in known volumes of 100% methanol, and used methods from Wada et al. (2007) to optimize use of enzyme immunoassay kits to measure cortisol. Then, we assayed selected EPFU hair samples across a range of sex, reproductive state, and ages (adult, juvenile) to explore the range of values in hair cortisol. Non-reproductive juvenile males had the highest mean cortisol levels (0.36 ng cortisol/mg hair) across sex, age and reproductive classes. Among all females, lactating females had the highest mean cortisol levels (0.22 ng/mg). Excluding pregnant females, hair cortisol did not vary with body mass, in either sex. Currently we are evaluating effects of sample time in freezer and of body location sampled on cortisol estimates.