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Frequency Encoding of Upconversion Nanoparticle Emission for Multiplexed Imaging of Spectrally and Spatially Overlapping Lanthanide Ions. | LitMetric
We present frequency encoded upconversion (FE-UPCON) widefield microscopy, an imaging approach that allows for multiplexed signal recovery based on frequency encoding of selected upconverted lanthanide ion emission rather than separation based on energy or time. FE-UPCON allows for the separation of luminescence from spectrally and spatially overlapping trivalent lanthanide ions (Ln) in upconversion nanoparticles (UCNPs). Utilizing the numerous electronic energy levels of Ln, one can generate a frequency encoded signal by periodic coexcitation with a secondary light source (modulated at a chosen frequency) that, for a particular wavelength, enhances the luminescence of the Ln of interest. We demonstrate that it is possible to selectively image spectrally overlapping UCNPs co-doped with Yb/Ho or Yb/Er by FE-UPCON in cells up to 10 frames per second on a conventional widefield microscope with the simple extension of an additional secondary light source and a chopper wheel for modulation. Additionally, we show that FE-UPCON does not compromise sensitivity and that single UCNP detection is obtainable. FE-UPCON adds a new dimension (frequency space) for multiplexed imaging with UCNPs.
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