Construction of FRET‐Based Off‐On Fluorescent Nanoprobes for Sensitive Detection of Intracellular Singlet Oxygen

Singlet oxygen (1O2) detection remains an active area of research in the field of biological and biomedical science. In this work, we reported a novel Off‐On type FRET‐based fluorescent 1O2 nanoprobe comprising the energy‐donating conjugated polymer, poly(9,9‐dioctylfluorenyl‐2,7‐diyl) (PFO), and the energy‐accepting 1O2 indicator, 3‐diphenylisobenzofuran (DPBF). Prior to the reaction with 1O2, the nanoprobe was in Off‐state because fluorescence of PFO (λem=441 nm) was completely quenched by loaded DPBF; in the presence of 1O2, it switched to On‐state due to the depletion of DPBF. The as‐prepared 1O2 nanoprobe had a particle size of ∼59 nm, good biocompatibility, high specificity and sensitivity with a limit of detection of 353 nM. Importantly, the fluorescent nanoprobes were highly sensitive to 1O2 under two‐photon excitation (λex=800 nm). Using a multiphoton confocal microscope, generation of intracellular 1O2 during photodynamic therapy was sensitively detected by recording fluorescence intensity of the nanoprobes. An Off‐On type FRET‐based fluorescent 1O2 nanoprobe was constructed from the energy‐donating conjugated polymer PFO and energy‐accepting 1O2 indicator DPBF. The nanoprobes exhibited good biocompatibility, high specificity and sensitivity. In the presence of 1O2, the fluorescent nanoprobe was switched from Off‐ to On‐state, which was employed to monitor the generation of intracellular 1O2 in real‐time during photodynamic therapy with 800 nm two‐photon excitation.