Abstract
Despite the potential of single-molecule sensing platforms to directly assay a multitude of different biomolecules, multiplexing single-molecule techniques to measure and classify many different molecules simultaneously remains challenging. The most common way to encode molecular identity in a single-molecule tag is to use color, but detector and signal-to-noise considerations limit the number of available color channels in most single-molecule instruments to at most 4-5 colors. However, additional spectroscopic information is present in fluorescence signals that can enable discrimination of chemically different fluorophores that have (for example) similar spectral emission but different brightness, fluorescence lifetime, or anisotropy.
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