Organelle-Targeting Polymer Dots Exhibiting Thermally Activated Delayed Fluorescence for Subcellular Imaging

Abstract

Selective imaging of specific subcellular structures provides valuable information about the cellular microenvironment. Materials exhibiting thermally activated delayed fluorescence (TADF) are rapidly emerging as metal-free probes with long-lived emission for intracellular time-gated imaging applications. Polymers incorporating TADF emitters can self-assemble into luminescent nanoparticles, termed polymer dots (Pdots), and this strategy enables them to circumvent the limitations of commercial organelle trackers and small molecule TADF emitters. In this study, diblock copolymers comprised of a hydrophilic block containing organelle-targeting monomers and a hydrophobic TADF-active block were synthesized by ringopening metathesis polymerization (ROMP). Oxanorbornene-based monomers incorporating morpholine and triphenylphosphonium groups for lysosome and mitochondria targeting, respectively, were also synthesized. ROMP by sequential addition yielded well-defined diblock copolymers with dispersities <1.28. To analyze the effect of tuning the hydrophilic corona on cellular viability and uptake, we prepared Pdots with poly(ethylene glycol) (PEG) and bis-guanidinium (BGN) coronas, resulting in limited and efficient cellular uptake, respectively. Red-emissive Pdots with BGN-based coronas and organelle-targeting functionality were obtained with quantum yields up to 12% in water under air. Colocalization analysis confirmed that lysosome and mitochondria labeling in live HeLa cells was accomplished within 2 h of incubation, affording Pearson's correlation coefficients of 0.37 and 0.70, respectively. The potential application of these Pdots for time-resolved imaging is highlighted by a proof of concept using time-gated spectroscopy, which effectively separates the delayed emission of the TADF Pdots from the background autofluorescence of biological serum.