Fluorescently Labeled Gold Nanoparticles with Minimal Fluorescence Quenching

Abstract

Minimal quenching of fluorescence emission was observed when the fluorescent dye AlexaFluor 514 (AF514) was covalently bound to gold monolayer protected clusters (AuMPCs) that have negligible plasmon bands (diameters <2 nm). The fluorescence emission of the conjugated dye was measured as a function of dye−AuMPC mixing ratio with a combination of steady-state and time-resolved ensemble spectroscopic measurements in conjunction with single-molecule fluorescence microscopy. Fluorescence emission of the conjugated samples decreased slightly as a function of dye mixing ratio, which, in combination with a negligible change in fluorescence lifetime, was attributed to static quenching of the dye by the AuMPCs. From the single-molecule fluorescence measurements, it was observed that luminescent conjugates could still be detected, and, at all loading ratios, almost all of the dye−particle conjugates photobleached in either a single- or double-step process, with a small subpopulation exhibiting more than two photobleaching events. Beyond simple, irreversible photobleaching, no additional blinking dynamics were detected at the single-molecule level within the time resolution of the experiment. Emission intensities of coupled fluorophores were comparable with those measured of dye molecules that had not been attached to the AuMPCs, suggesting that the statically quenched fluorophores are entirely nonemissive, whereas the remaining dyes are essentially unquenched. These results are discussed in the context of other dye−AuMPC coupled systems described in the literature.