Determining Essential\nRequirements for Fluorophore\nSelection in Various Fluorescence Applications Taking Advantage of\nDiverse Structure–Fluorescence Information of Chromone Derivatives

Herein,\nwe report our work exploring the essential requirements\nfor fluorophore selection during the development of various fluorescence\napplications. We assembled a library of chromone-derived fluorophores\nwith diverse structure–fluorescence properties, which allowed\nus to choose the fluorophore pairs with similar structures but differing\nfluorescence properties and compared the performance of the selected\nfluorophore pairs in three types of commonly used fluorescence applications.\nWe found that the selection standard of a suitable fluorophore is\nvariable depending on the application. (1) In fluorescence imaging,\nfluorophores with strong and constant fluorescence under various conditions,\nsuch as a large pH range, are preferred. Notably, (2) in the detection\nof bioactive species, fluorophores with relatively lower fluorescence\nquantum yield favor the detection sensitivity. Furthermore, (3) in\nenzymatic assays employing fluorescence, the key parameter is the\nbinding affinity between the fluorophore and the enzyme.