Vitamin D is an essential hormone in vertebrates, and these animals can acquire vitamin D through their diet, ultraviolet B (UVB) radiation, or both. It is important to identify which route a species utilizes. For example, cats and dogs only acquire vitamin D through their diets, while humans acquire vitamin D through both routes. In reptiles, most studies have focused on evaluating the effects of UVB exposure on vitamin D status, with limited study into the impacts of diet. The goal of this study was to measure the plasma 25-hydroxyvitamin D3 (25-OHD3) concentrations of northern blue tongued skinks (Tiliqua scincoides intermedia) fed wet cat food provided by the breeder (Wellness®, Burlington, MA, USA) and not exposed to UVB. The authors’ hypothesis for this study was that the skinks would have 25-OHD3 concentrations > 50 nmol/L because of the vitamin D3 supplemented cat food (573 IU/kg, as fed). Eleven yearling skinks were used for this cross-sectional study. The skinks were captive bred and raised on the commercial cat food over the past year. Blood samples were collected from the ventral tail/jugular vein, placed in lithium heparin microtainers, centrifuged, and the plasma frozen at -80oC. The samples were transported to the Michigan State University Diagnostic Laboratory for radioimmunoassay testing. The 25-OHD3 concentrations were normally distributed: 17.1±3.8 nmol/L, min-max: 9-22. These concentrations were lower than expected, and analogous to 25-OHD3 concentrations in reptiles not exposed to UVB. Based on these results, the authors recommend a follow-up study measuring 25-OHD3 concentrations post-UVB exposure.