Shortwave infrared fluorofluorophores for multicolor in vivo imaging.

Developing chemical tools to detect and influence biological processes is a cornerstone of chemical biology. Here we combine two tools which rely on orthogonality- perfluorocarbons and multiplexed shortwave infrared fluorescence imaging- to track the biodistribution of nanoemulsions in real time in living mice. Drawing inspiration from fluorous and shortwave infrared (SWIR) fluorophore development, we prepared two SWIR-emissive, fluorous-soluble chromenylium polymethine dyes. These are the most red-shifted fluorous fluorophores- "fluorofluorophores"- to date. After characterizing the dyes, their utility was demonstrated by tracking perfluorocarbon nanoemulsion biodistribution in vivo. Using an excitation-multiplexed approach to visualize two variables simultaneously, we gained insight into the importance of size and surfactant identity on biodistribution.