Two-photon excitation fluorescent spectral and decay properties of retrograde neuronal tracer Fluoro-Gold.
Two-photon excitation fluorescent spectral and decay properties of retrograde neuronal tracer Fluoro-Gold.
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DOI：
10.1038/s41598-021-97562-3
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发表时间：
2021-09-10
影响因子：
4.6
通讯作者：
Jowett N
中科院分区：
文献类型：
--
作者：
Miller MQ;Hernández IC;Chacko JV;Minderler S;Jowett N
荧光金是一种荧光神经元示踪剂，适用于神经系统的靶向深度成像。宽视场荧光显微镜能够显示荧光金，但缺乏深度分辨能力。虽然扫描激光共聚焦显微镜可以获得体积数据，但成像深度是有限的，而最佳的荧光金的单光子激发需要一条非传统的紫外光激发线。双光子激发显微镜使用超快脉冲红外激光在不透明组织中以无与伦比的深度以高分辨率成像荧光团。荧光金标记神经元的深度成像具有促进对中枢和外周神经系统的理解的潜力，但其双光子光谱和时间特性仍未得到表征。在这里，我们报道了荧光金在720-990 nm之间的双光子激发光谱，以及它在水溶液和小鼠脑干组织中的荧光衰减率。我们通过荧光金标记的面部运动核团的双光子激发显微镜展示了前所未有的整体安装的小鼠脑干的成像深度。在显微镜调谐范围内，荧光金的最佳双光子激发出现在720 nm处，最大寿命对比度出现在760 nm处，平均荧光寿命为1.4 ns。通过浸泡在折射率匹配溶液中，整个安装的脑干外植体可以很容易地成像到超过450微米的深度。
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.
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影响因子：
4.6
作者：
Lee E;Choi J;Jo Y;Kim JY;Jang YJ;Lee HM;Kim SY;Lee HJ;Cho K;Jung N;Hur EM;Jeong SJ;Moon C;Choe Y;Rhyu IJ;Kim H;Sun W
通讯作者：
Sun W
影响因子：
2.9
作者：
AKINTUNDE, A;BUXTON, DF
通讯作者：
BUXTON, DF
影响因子：
0.8
作者：
Drobizhev, Mikhail;Molina, Rosana S.;Hughes, Thomas E.
通讯作者：
Hughes, Thomas E.
影响因子：
--
作者：
Grant, Gunnar
通讯作者：
Grant, Gunnar
影响因子：
64.8
作者：
通讯作者：
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