Two-photon excitation fluorescent spectral and decay properties of retrograde neuronal tracer Fluoro-Gold.

Autor: Miller MQ; Surgical Photonics and Engineering Laboratory, Massachusetts Eye and Ear, Harvard Medical School, 243 Charles Street, Boston, MA, 02114, USA.; Department of Otolaryngology/Head and Neck Surgery, University of North Carolina Health Care, Chapel Hill, NC, USA., Hernández IC; Surgical Photonics and Engineering Laboratory, Massachusetts Eye and Ear, Harvard Medical School, 243 Charles Street, Boston, MA, 02114, USA. ivan_cotohernandez@meei.harvard.edu., Chacko JV; Laboratory for Optical and Computational Instrumentation, University of Wisconsin, Madison, WI, USA., Minderler S; Surgical Photonics and Engineering Laboratory, Massachusetts Eye and Ear, Harvard Medical School, 243 Charles Street, Boston, MA, 02114, USA., Jowett N; Surgical Photonics and Engineering Laboratory, Massachusetts Eye and Ear, Harvard Medical School, 243 Charles Street, Boston, MA, 02114, USA. nate_jowett@meei.harvard.edu.
Jazyk: angličtina
Zdroj: Scientific reports [Sci Rep] 2021 Sep 10; Vol. 11 (1), pp. 18053. Date of Electronic Publication: 2021 Sep 10.
DOI: 10.1038/s41598-021-97562-3
Abstrakt: Fluoro-Gold is a fluorescent neuronal tracer suitable for targeted deep imaging of the nervous system. Widefield fluorescence microscopy enables visualization of Fluoro-Gold, but lacks depth discrimination. Though scanning laser confocal microscopy yields volumetric data, imaging depth is limited, and optimal single-photon excitation of Fluoro-Gold requires an unconventional ultraviolet excitation line. Two-photon excitation microscopy employs ultrafast pulsed infrared lasers to image fluorophores at high-resolution at unparalleled depths in opaque tissue. Deep imaging of Fluoro-Gold-labeled neurons carries potential to advance understanding of the central and peripheral nervous systems, yet its two-photon spectral and temporal properties remain uncharacterized. Herein, we report the two-photon excitation spectrum of Fluoro-Gold between 720 and 990 nm, and its fluorescence decay rate in aqueous solution and murine brainstem tissue. We demonstrate unprecedented imaging depth of whole-mounted murine brainstem via two-photon excitation microscopy of Fluoro-Gold labeled facial motor nuclei. Optimal two-photon excitation of Fluoro-Gold within microscope tuning range occurred at 720 nm, while maximum lifetime contrast was observed at 760 nm with mean fluorescence lifetime of 1.4 ns. Whole-mount brainstem explants were readily imaged to depths in excess of 450 µm via immersion in refractive-index matching solution.
(© 2021. The Author(s).)
Databáze: MEDLINE
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