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© 2021 American Chemical Society.It is well–known that plasmonic nanoparticles can modify the spectroscopic properties of nearby optical probes, for example, enhanced emission of a fluorescent dye. Yet, the detection and quantification of this effect in bulk solution remain challenging even while size– and shape–controlled nanoparticles have become readily available. We investigated this problem and identified two main difficulties which we were able to overcome through systematic studies. For the detection of fluorescence emanating from optically dense nanoparticle solutions, we describe an analytical model that provides guidelines for experimentalists to maximize the fluorescence intensity by optimizing the concentration, light paths, and excitation–detection volume of the sample. For the quantification of enhancement, which critically hinges upon the comparison to an accurate reference sample, we exploit the tools of DNA nanotechnology to remove the fluorophore from plasmonic coupling on–demand, forming an in situ reference. Using a model system of fluorophore Cy3 and 80 nm gold nanoparticles, we show that these strategies enable the quantitative measurement of plasmonic enhancement across a 20–fold range of optical densities. We anticipate that the presented experimental framework will allow for routine, quantitative measurements for the research and development of plasmon–enhanced phenomena.
