Optical imaging within the shortwave infrared (SWIR, 1000−2000 nm) region of the electromagnetic spectrum has enabled high-resolution and high-contrast imaging in mice, non-invasively. Polymethine dyes, with their narrow absorption spectra and high absorption coefficients, are optimal probes for fast and multiplexed SWIR imaging. Here, we expand upon the multiplexing capabilities in SWIR imaging by obtaining brighter polymethine dyes with varied excitation wavelengths spaced throughout the near-infrared (700−1000 nm) region. Building on the flavylium polymethine dye scaffold, we explored derivatives with functional group substitution at the 2-position, deemed chromenylium polymethine dyes. The reported dyes have reduced nonradiative rates and enhanced emissive properties, enabling noninvasive imaging in mice in a single color at 300 fps and in three colors at 100 fps. Combined with polymethine dyes containing a red-shifted julolidine flavylium heterocycle and indocyanine green, distinct channels with well-separated excitation wavelengths provide non-invasive video-rate in vivo imaging in four colors.

The bioorthogonal nature of perfluorocarbons provides a unique platform for introducing dynamic nano-and microdroplets into cells and organisms. To monitor the localization and deformation of the droplets, fluorous soluble fluorophores that are compatible with standard fluorescent protein markers and applicable to cells, tissues, and small organisms are necessary. Here, we introduce fluorous cyanine dyes that represent the most red-shifted fluorous soluble fluorophores to date. We study the effect of covalently appended fluorous tags on the cyanine scaffold and evaluate the changes in photophysical properties imparted by the fluorous phase. Ultimately, we showcase the utility of the fluorous soluble pentamethine cyanine dye for tracking the localization of perfluorocarbon nanoemulsions in macrophage cells and for measurements of mechanical forces in multicellular spheroids and zebrafish embryonic tissues. These studies demonstrate that the red shifted cyanine dyes offer spectral flexibility in multiplexed imaging experiments and enhanced precision in force measurements.

With the growing development of new contrast agents for optical imaging using near‐infrared and shortwave infrared (SWIR) wavelengths, it is essential to have consistent benchmarks for emitters in these regions. Indocyanine green (ICG), a ubiquitous and FDA‐approved organic dye and optical imaging agent, is commonly employed as a standard for photophysical properties and biological performance for imaging experiments at these wavelengths. Yet, its reported photophysical properties across organic and aqueous solvents vary greatly in the literature, which hinders its ability to be used as a consistent benchmark. Herein, we measure photophysical properties in organic and aqueous solvents using InGaAs detection (∼950–1,700 nm), providing particular relevance for SWIR imaging.

Photo-unclick chemistry mediates visible and near IR-controlled drug release via a singlet oxygen (SO)-cleavable linker. Due to the limited diffusion distance of SO in biological systems, a photosensitizer and the SO-cleavable linker have been conjugated in one molecule or mixed in nano-drug delivery systems. In this communication, we demonstrate a new strategy to activate prodrugs with photo-unclick chemistry in an intermolecular fashion using an SO-cleavable CA4 prodrug and a mitochondria-specific photosensitizer, protoporphyrin IX, formed from prodrug hexyl-5-aminolevulinate.