The β-diketone moiety is commonly present in many anticancer drugs, antibiotics, and natural products. We describe a general method for radiolabeling β-diketone-bearing molecules with fluoride-18. Radiolabeling is carried out via (18)F-(19)F isotopic exchange on non-radioactive difluoro-dioxaborinins, which are generated by minimally modifying the β-diketone as a difluoroborate. Radiochemistry is one-step, rapid (< 10 min), high-yielding (> 80%), and proceeds at room temperature to accommodate the half-life of F-18 (t(1/2) = 110 min). High molar activities (7.4 Ci/μmol) were achieved with relatively low starting activities (16.4 mCi). It was found that substituents affect both the solvolytic stability and fluorescence properties of difluoro-dioxaborinins. An F-18 radiolabeled difluoro-dioxaborinin probe that is simultaneously fluorescent showed sufficient stability for in vivo PET/fluorescence imaging in mice, rabbits, and patients. These findings will guide: the design of probes with specific PET/fluorescence properties; the development of new PET/fluorescence dual-modality reporters; and accurate in vivo tracking of β-diketone molecules.