Laurell F. Kessler, Ashwin Balakrishnan, Tanja Menche, Dongni Wang, Yunqing Li, Maximilian Mantel, Marius Glogger, Marina S. Dietz and Mike Heilemann*,
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引用次数: 0
摘要
通过荧光团标记的 DNA 短寡核苷酸的瞬时和重复杂交进行蛋白质标记,已广泛应用于各种光学超分辨率显微镜方法。其主要优点是多目标成像和分子定量。但溶液中存在未结合的荧光团-DNA 标记会产生高背景信号,这是一个挑战。在此,我们报告了荧光团二聚体与 DNA 寡核苷酸共轭的自淬灭方法,这是减少荧光背景的一般概念。杂交时,两种荧光团的荧光信号都会恢复。我们扩展了适合自淬灭的荧光团工具箱,并报告了它们的光谱和杂交平衡。我们将自淬火荧光团-DNA 标签应用于受激发射耗尽显微镜和单分子定位显微镜,并报告了改进的成像性能。
Self-Quenched Fluorophore-DNA Labels for Super-Resolution Fluorescence Microscopy
Protein labeling through transient and repetitive hybridization of short, fluorophore-labeled DNA oligonucleotides has become widely applied in various optical super-resolution microscopy methods. The main advantages are multitarget imaging and molecular quantification. A challenge is the high background signal originating from the presence of unbound fluorophore-DNA labels in solution. Here, we report the self-quenching of fluorophore dimers conjugated to DNA oligonucleotides as a general concept to reduce the fluorescence background. Upon hybridization, the fluorescence signals of both fluorophores are restored. We expand the toolbox of fluorophores suitable for self-quenching and report their spectra and hybridization equilibria. We apply self-quenched fluorophore-DNA labels to stimulated emission depletion microscopy and single-molecule localization microscopy and report improved imaging performances.
期刊介绍:
An essential criterion for acceptance of research articles in the journal is that they provide new physical insight. Please refer to the New Physical Insights virtual issue on what constitutes new physical insight. Manuscripts that are essentially reporting data or applications of data are, in general, not suitable for publication in JPC B.