Bathochromic Shift via C=O to C=S Substitution: A Far-Red Fluorogen for Multiplexed FLIM with FAST Fluorogen-Activating Protein
The palette of the fluorogen-activating protein FAST expanded into the far-red region by the development of a novel fluorogen, <b>HBTR-3,5-DOM</b>. 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 <b>HBTR-3,5-DOM</b> 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 using fluorescence lifetime imaging microscopy (FLIM).
Authors
- Михаил С. Баранов (ORCID: https://orcid.org/0000-0002-9339-7603)
- Aidar R. Gilvanov (ORCID: https://orcid.org/0000-0001-6174-7798)
- Marina V. Goncharuk (ORCID: https://orcid.org/0000-0002-6984-7125)
- Andrey А. Mikhaylov (ORCID: https://orcid.org/0000-0001-9433-9141)
- Svetlana V. Sidorenko (ORCID: https://orcid.org/0000-0002-4074-0500)
- Sergey A. Goncharuk (ORCID: https://orcid.org/0000-0002-0263-6462)
- Eugene G. Maksimov (ORCID: https://orcid.org/0000-0001-6767-1501)
- Artur V. Eshtukov‐Shcheglov
- Marina V. Molchanova (ORCID: https://orcid.org/0009-0007-5685-5138)
- S. A. Krasnova (ORCID: https://orcid.org/0009-0002-7401-1928)
- Yulia A. Bogdanova (ORCID: https://orcid.org/0000-0003-1745-4204)
Institutions
- Lomonosov Moscow State University (RU)
- Pirogov Russian National Research Medical University (RU)
- Institute of Bioorganic Chemistry (RU)
Publication Details
- Journal
- International Journal of Molecular Sciences
- Published
- 2025-12-19
- DOI
- https://doi.org/10.3390/ijms27010023
- Primary Topic
- Advanced Fluorescence Microscopy Techniques
- Type
- article
- Field-Weighted Citation Impact
- 0.00