Kinetically stable sub-50 nm fluorescent block copolymer nanoparticles via photomediated RAFT dispersion polymerization for cellular imaging

Self-assembled block copolymer nanoparticles (NPs) have emerged as major potential nanoscale vehicles for fluorescence bioimaging. The preparation of NPs with high yields possessing high kinetic stability to prevent the leakage of fluorophore molecules is crucial to their practical implementation. Here, we report a photomediated RAFT polymerization-induced self-assembly (PISA) yielding uniform and nanosized poly((oligo(ethylene glycol) acrylate)-block-poly(benzyl acrylate) particles (POEGA-b-PBzA) with a concentration of 22 wt%, over 20 times more than with micellization and nanoprecipitation. The spherical diblock copolymer nanoparticles have an average size of 10-50 nm controllable through the degree of polymerization of the stabilizing POEGA block. Subsequent dialysis against water and swelling with Nile red solution led to highly stable fluorescent NPs able to withstand the changes in concentration, ionic strength, pH or temperature. A PBzA/water interfacial tension of 48.6 mN m-1 hinders the exchange between copolymer chains, resulting in the trapping of NPs in a "kinetically frozen" state responsible for high stability. A spectroscopic study combining fluorescence and UV-vis absorption agrees with a preferential distribution of fluorophores in the outer POEGEA shell despite its hydrophobic nature. Nile red-doped POEGA-b-PBzA micelles without initiator residues and unimers but with high structural stability turn out to be noncytotoxic, and can be used for the optical imaging of cells. Real-time confocal fluorescence microscopy shows a fast cellular uptake using C2C12 cell lines in minutes, and a preferential localization in the perinuclear region, in particular in the vesicles.