Vitamin D is an essential hormone that can be acquired via the diet, exposure to ultraviolet B (UVB) radiation, or a combination of both. Studies in reptiles suggest that the acquisition of vitamin D can vary between species; thus, species-specific evidence-based research should be pursued to develop appropriate husbandry recommendations. The objective of this study was to determine whether artificial UVB could be used to increase circulating 25-hydroxyvitamin D3 (25-OHD3) concentrations in juvenile Blanding's turtles (Emydoidea blandingii). Sixteen juvenile Blanding's turtles from an ongoing headstart program at the DuPage County Forest Preservation District (Wheaton, IL, USA) were used for this study. The Blanding's turtles were randomly divided into two groups of eight by using a random number generator. The treatment group was exposed to light from three compact UVB 23-W fluorescent bulbs 12 h/day for 6 months, whereas the control group was not provided supplemental UVB lighting. Bulbs were placed 15.2 cm (6 in) from the water surface. A radiometer was used to measure UVB radiation 15.2 cm from the bulb surface on days 0 and 180. Blood samples were collected at a single time point (day 180) to measure 25-OHD3 concentrations. There was a significant difference (P < 0.001) in plasma 25-OHD3 concentrations between groups, with 25-OHD3 concentrations being 5.5 times higher in the UVB group than in the controls. Based on the results of this study, we recommend exposing juvenile Blanding's turtles to artificial UVB as part of their standard care.