The aim of this study is to determine if ultraviolet light (UVC) irradiation in combination with fluorescence-guided surgery (FGS) can eradicate metastatic human pancreatic cancer in orthotopic nude-mouse models. Two weeks after orthotopic implantation of human MiaPaCa-2 pancreatic cancer cells, expressing green fluorescent protein (GFP), in nude mice, bright-light surgery (BLS) was performed on all tumor-bearing mice (n = 24). After BLS, mice were randomized into 3 treatment groups; BLS-only (n = 8) or FGS (n = 8) or FGS-UVC (n = 8). The residual tumors were resected using a hand-held portable imaging system under fluorescence navigation in mice treated with FGS and FGS-UVC. The surgical resection bed was irradiated with 2700 J/m2 UVC (254 nm) in the mice treated with FGS-UVC. The average residual tumor area after FGS (n = 16) was significantly smaller than after BLS only (n = 24) (0.135±0.137 mm2 and 3.338±2.929 mm2, respectively; p = 0.007). The BLS treated mice had significantly reduced survival compared to FGS- and FGS-UVC-treated mice for both relapse-free survival (RFS) (p<0.001 and p<0.001, respectively) and overall survival (OS) (p<0.001 and p<0.001, respectively). FGS-UVC-treated mice had increased RFS and OS compared to FGS-only treated mice (p = 0.008 and p = 0.025, respectively); with RFS lasting at least 150 days indicating the animals were cured. The results of the present study suggest that UVC irradiation in combination with FGS has clinical potential to increase survival.