Experimental validation of eosin-mediated photo-redox polymerization mechanism and implications for signal amplification applications

Abstract

Oxygen-tolerant radical polymerization has demonstrated applications in biosensors as a signal amplification method for molecular recognition events. In particular, eosin-mediated photo-redox polymerization, a visible light-initiated radical copolymerization method using N-vinyl pyrrolidone and PEDGA monomers, can be performed in aqueous microliter-scale droplets under atmospheric conditions, and has been used for rapid (≤90 s) signal amplification in several diagnostic assays. In recent years, significant progress has been made in understanding the reaction mechanism, and here we assess the accuracy of the proposed mechanism via experimental validation in an assay format. A 2D reaction–diffusion model was developed and compared to experimental behavior of eosin photopolymerization for paper-based signal amplification. For 3 and 4 mm test zones, the model predicted, within an order of magnitude, experimentally observed effects of oxygen exposure and eosin photoinitiator concentration on polymer formation in a droplet. Both model and experimental results demonstrated that high oxygen exposure and low eosin concentration restrict polymer formation to the center of circular wells. Decreasing the surface-area-to-volume ratio of the reaction droplet and increasing eosin concentration allow polymerization throughout the zone, initially forming in radially intermediate zones due to oxygen's role not just as a reaction inhibitor but also a promoter via photoinitiator regeneration. Reaction volumes as low as 20 μL on 3, 4, and 5 mm diameter reaction zones enabled sensitive signal amplification, although higher oxygen exposure (3–10 μL droplets) showed greatly reduced sensitivity. Observing oxygen tolerance limits and experimentally validating the reaction mechanism can help better understand the eosin photopolymerization system and its applications in diagnostic assay signal amplification.