Heat shock proteins (HSPs) are important mediators of the normal cellular function and the cellular stress response. As such, HSPs are often utilized to measure the effects of stressors on organisms in vivo. However, multiple variables can influence their expression, including time or season, confounding results. To investigate the utility of HSPs in measuring effects of stressors in a top trophic carnivore, we captured 20 American alligators, Alligator mississipiensis (Daudin, 1802), placed them in burlap sacks for 2 hours and collected blood samples over four time points (baseline, 1 and 2 hours after placement into burlap sacks, and 24 hours after initial capture) to measure plasma corticosterone (the main crocodilian glucocorticoid) and levels of HSP60, HSP70, and HSP90. Time point significantly affected plasma corticosterone levels in alligators (p < 0.0001), with levels significantly elevated at 1, 2, and 24 hours, relative to baseline (all p < 0.05). However, capture stress did not affect HSP60, HSP70, or HSP90 in red blood cells (all p > 0.05). Our results suggest HSPs may be important biomarkers for investigating the impacts of stressors on captive and wild crocodilians, since they are not acutely elevated by capture or handling stress.