Mechanistic Insights into Polymerization-Induced Self-Assembly Using Maleimide-Based Fluorophores

Polymerization-induced self-assembly (PISA) is a versatile and readily accessible method to produce nanoparticles of various morphologies in situ as polymerization progresses. PISA exploits the chain extension of a solvophilic macromolecular chain-transfer agent with monomers that are miscible in the continuous phase but form a solvophobic, immiscible polymer, driving self-assembly. However, the ability to monitor in situ the onset of self-assembly and the evolution of morphology during the PISA process remains a significant challenge, which critically limits our understanding of the mechanisms of particle formation. In this work, we demonstrate that a maleimide-based small-molecule fluorophore can act as a powerful probe to study PISA over time using fluorescence and fluorescence lifetime as outputs. We show that the aminochloromaleimide (ACM) fluorophore can be readily incorporated within a PISA system to produce fluorescent nanostructures without affecting their final morphology in comparison to their nonfluorescent analogues. The ACM probe exhibits diagnostic increases in fluorescence lifetime first with the onset of self-assembly and then with the evolution of particle morphology in the order of spheres > vesicles > worms. Excitingly, monitoring the change in fluorescence lifetime in situ during PISA yielded insights into the mechanism of particle formation when targeting higher-order morphologies. Finally, we demonstrate that these maleimide-functionalized nanostructures can be used as cell imaging agents using fluorescence lifetime imaging microscopy (FLIM), whereby each morphology produces distinct lifetime decay patterns within a cell environment. Overall, we envision this becoming a powerful tool for the analysis of nanoparticle states within complex environments, inspiring further investigations of the study of PISA using this simple and accessible method.