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Single-chromophore-based photoswitchable nanoparticles enable dual-alternating-color fluorescence for unambiguous live cell imaging.

We have developed a class of spiropyran dyes and their fluorescence colors can be reversibly photoswitched from red to green, blue, or nearly dark, thus alternating between two colors. Such individual dyes emit either one color or the other but not both simultaneously. Nanoparticles enabled with these photoswitchable dyes, however, emit either one pure color or a combination of both colors because the nanoparticle fluorescence originates from multiple dyes therein. As a result, the nanoparticle shines >30 times brighter than state-of-the-art organic dyes such as fluorescein. Interestingly, these copolymer nanoparticles exhibit tunable nonspecific interactions with live cells, and nanoparticles containing properly balanced butyl acrylate and acrylamide monomers render essentially very little nonspecific binding to live cells. Decorated with HMGA1 protein, these optically switchable dual-color nanoparticles undergo endocytosis and unambiguously identify themselves from fluorescence interference including autofluorescence, thus enabling a new tool for live cell imaging.