Multicolored pH-Tunable and Activatable Fluorescence Nanoplatform Responsive to Physiologic pH Stimuli

Tunable, ultra-pH responsive fluorescent nanoparticles with multichromatic emissions are highly valuable in a variety of biological studies, such as endocytic trafficking, endosome/lysosome maturation, and pH regulation in subcellular organelles. Small differences (e.g., <1 pH unit) and yet finely regulated physiological pH inside different endocytic compartments present a huge challenge to the design of such a system. Herein, we report a general strategy to produce pH-tunable, highly activatable multicolored fluorescent nanoparticles using commonly available pH-insensitive dyes with emission wavelengths from green to near IR range. The primary driving force of fluorescence activation between the ON (unimer) and OFF (micelle) states is the pH-induced micellization. Among three possible photochemical mechanisms, homo Förster resonance energy transfer (homoFRET)-enhanced decay was found to be the most facile strategy to render ultra-pH response over the H-dimer and photoinduced electron transfer (PeT) mechanisms. Based on this insight, we selected several fluorophores with small Stoke shifts (<40 nm) and established a panel of multicolored nanoparticles with wide emission range (500-820 nm) and different pH transitions. Each nanoparticle maintained the sharp pH response (ON/OFF < 0.25 pH unit) with corresponding pH transition point at pH 5.2, 6.4, 6.9, and 7.2. Incubation of a mixture of multicolored nanoparticles with human H2009 lung cancer cells demonstrated sequential activation of the nanoparticles inside endocytic compartments directly correlating with their pH transitions. This multicolored, pH-tunable nanoplatform offers exciting opportunities for the study of many important cell physiological processes, such as pH regulation and endocytic trafficking of subcellular organelles.