Color-tuned FRET polystyrene microspheres by single wavelength excitation

Fluorescent monodisperse polystyrene microspheres were prepared by two-stage dispersion polymerization, which successfully covalently labeled microspheres with two dyes without disturbing the final particle size and size distribution. By varying the dye concentrations, microspheres show tuned colors with different fluorescent intensity under a single wavelength excitation. Fluorescence resonance energy transfer (FRET) between two labeled dyes was proved to contribute to the emission of the longer-wavelength dye at a shorter-wavelength excitation. There is no dye leakage for microspheres because of the covalent incorporation of dye molecules. The microsphere matrix provides good protection of dye molecules and blocks the influence of media outside on the fluorescence of microspheres. Single microsphere shows intense fluorescence due to a large number of encapsulated dye molecules. These uniform barcoding fluorescent microspheres have potential application in multiplexed bioanalysis.