Bathochromic Shift via C=O to C=S Substitution: A Far-Red Fluorogen for Multiplexed FLIM with FAST Fluorogen-Activating Protein.

ABSTRACT: The palette of the fluorogen-activating protein FAST expanded into the far-red region by the development of a novel fluorogen, HBTR-3,5-DOM. This was achieved through a C=O to C=S substitution in the classic hydroxybenzylidene-rhodanine core, which induced a bathochromic shift of over 100 nm. The complexes of HBTR-3,5-DOM with FAST variants pFAST and F62L are characterized by absorption and emission maxima at 640-650 nm and ~670 nm, respectively, and are found to exhibit distinct fluorescence lifetimes. The fluorogen is successfully applied in genetically encoded live-cell imaging together with these FAST variants for various subcellular structures. Furthermore, its potential for multiplexed imaging is demonstrated by the simultaneous discrimination of two targeted proteins u

SUBMITTER: Gilvanov AR 

PROVIDER: S-EPMC12786233 | biostudies-literature | 2025 Dec

REPOSITORIES: biostudies-literature