<p>Conventional dark-ink tattoos used for radiotherapy positioning may remain permanently visible. We developed a fluorescent tattoo system comprising a medical tattoo device, fluorescent ink, and ultraviolet (UV) lamps and evaluated its clinical feasibility. Twenty-four patients prospectively received fluorescent tattoos, and setup displacement and procedure times were compared with those of 24 retrospectively selected patients with conventional dark-ink tattoos. The time required for tattooing and patient treatment setup was estimated. The setup errors were determined using CBCT. Pain, satisfaction, and adverse events were evaluated prospectively in the fluorescent group. Lamps with wavelengths in the UVA range were developed. The appropriate needle vibration frequency and needle type for tattoo procedures were determined. CBCT-based setup displacement did not differ significantly between groups. Median CT simulation time was longer with fluorescent tattoos, whereas treatment setup time was similar. The VAS score was evaluated as 1.9 ± 0.8. The patients were highly satisfied with the availability of showers and the invisible marker. No tattoo-related adverse events were reported during a three-year medical-record review. In this small, non-randomized cohort, the fluorescent tattoo system was clinically feasible and showed no significant difference in CBCT-based setup displacement from conventional dark-ink tattoos. It may offer a cosmetically inconspicuous option for selected patients, although UV illumination, longer application time, limited cohort diversity, and long-term pigment safety require further study.</p>