Evaluating resource use patterns for imperiled species is critical for understanding what supports their populations. Here we established stable isotope (δ13C, δ15N) values for the endangered green sea turtle (Chelonia mydas) population found within the boundaries of Dry Tortugas National Park (DRTO), south Florida, USA. There is little gene flow between turtles sampled at DRTO and in other rookeries in Florida, underscoring the need to study this distinct population. Between 2008 and 2015 we collected multiple sample types (skin [homogenized epidermis/dermis], whole blood, red blood cells, plasma, carapace) from 151 unique green turtles, including 43 nesting females and 108 in-water captures; some individuals were resampled multiple times across years to evaluate consistency of isotope signatures. Isotopic ratios ranged from -27.3 to -5.4 for δ¹³C and 3.7 to 10.6 for δ¹⁵N. Using linear mixed models, we evaluated covariates (sample type, turtle size and year) that best explained the isotope patterns observed in turtle tissues. Predictions from the top model for δ¹³C indicated a slight decrease over time and for δ¹⁵N a slight increase in the middle sampling years (2010–2012); results indicated that turtle size appeared to be the driver behind the range in δ¹³C and δ¹⁵N observed in turtle skin. We found a pattern in stable carbon isotope values that are indicative of an ontogenetic change from an omnivorous diet in smaller turtles to a seagrass-based diet in larger turtles. When we compared the stable carbon and nitrogen isotope values of the samples collected from turtles with that of seagrasses found in DRTO, we found that turtles > 65 cm SCL had similar stable carbon isotope values to the seagrass species present. Results of this study suggest stable isotope analysis coupled with data for available resources can be useful for tracking and detecting future changes in green turtle resource shifts in DRTO.