Frequency-encoded multicolor fluorescence imaging with single-photon counting color-blind detection

Standard fluorescence microscopy relies on filter-based detection of emitted photons after fluorophore excitation at the appropriate wavelength. While of enormous utility to the biological community, the implementation of approaches for simultaneous multicolor fluorescence imaging is commonly challenged by the large spectral overlap between different fluorophores. In here we describe an alternative multicolor fluorescence imaging methodology that exclusively relies on the absorption spectra of the fluorophores instead of their fluorescence emissions.

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