Red-Emitting Rhodamine Dyes for Fluorescence Microscopy and Nanoscopy
Red-Emitting Rhodamine Dyes for Fluorescence Microscopy and Nanoscopy
复制标题
DOI：
10.1002/chem.200902309
复制
发表时间：
2010-01-01
影响因子：
4.3
通讯作者：
Hell, Stefan W.
中科院分区：
文献类型：
--
作者：
Kolmakov, Kirill;Belov, Vladimir N.;Hell, Stefan W.
荧光标记在生物显微镜中是非常有价值的，因为它们最大限度地减少了细胞自身荧光，增加了多色实验的灵活性。研制了一种新型罗丹明染料，可被630 nm激光激发，发射波长在660 nm左右。新型罗丹明具有很强的光稳定性，荧光量子产率高达80%。激发态寿命长3.4 ns，系统间交叉率相对较低。它们在传统和亚衍射分辨率显微镜(如STED（受激发射损耗）和GSDIM（基态损耗与单个分子返回）以及基于单分子的实验（如荧光相关光谱（FCS））中都表现良好。描述了从同一含发色团支架开始的亲脂和亲水衍生物的合成。引入两个亚砜基团提供了在水中的高溶解度和荧光量子产率的显著提高。氨基或硫醇反应基团的附着使染料在生物学中用作荧光标记。在某些位置氘化的染料具有狭窄和对称的分子质量分布模式，并被提出作为MS或LC-MS的新标签，用于复杂混合物中各种物质类别（如胺和硫醇）的鉴定和定量。高分辨率GSDIM图像和标记微管和脂质的活细胞STED-FCS实验证明了新型探针在现代荧光显微镜和纳米显微镜中的多功能性。
Fluorescent markers emitting in the red are extremely valuable in biological microscopy since they minimize cellular autofluorescence and increase flexibility in multicolor experiments. Novel rhodamine dyes excitable with 630 nm laser light and emitting at around 660 nm have been developed. The new rhodamines are very photostable and have high fluorescence quantum yields of up to 80%. long excited state lifetimes of 3.4 ns, and comparatively low intersystem-crossing rates. They perform very well both in conventional and in subdiffraction-resolution microscopy such as STED (stimulated emission depletion) and GSDIM (ground-state depletion with individual molecular return), as well as in single-molecule-based experiments such as fluorescence correlation spectroscopy (FCS). Syntheses of lipophilic and hydrophilic derivatives starting from the same chromophore-containing scaffold are described. Introduction of two sulfo groups provides high solubility in water and a considerable rise in fluorescence quantum yield. The attachment of amino or thiol reactive groups allows the dyes to be used as fluorescent markers in biology. Dyes deuterated at certain positions have narrow and symmetrical molecular mass distribution patterns, and are proposed as new tags in MS or LC-MS for identification and quantification of various substance classes (e.g., amines and thiols) in complex mixtures. High-resolution GSDIM images and live-cell STED-FCS experiments on labeled microtubules and lipids prove the versatility of the novel probes for modern fluorescence microscopy and nanoscopy.