A Bright Surprise: Live-Cell Labeling with Negatively Charged Fluorescent Probes based on Disulfonated Rhodamines and HaloTag

IF 6.1 Q1 CHEMISTRY, MULTIDISCIPLINARY Chemistry methods : new approaches to solving problems in chemistry Pub Date : 2023-04-13 DOI:10.1002/cmtd.202200076

Dr. Dojin Kim, Dr. Stefan Stoldt, Dr. Michael Weber, Prof. Stefan Jakobs, Dr. Vladimir N. Belov, Prof. Stefan W. Hell

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引用次数: 3

Abstract

Disulfonated rhodamines are photostable and bright dyes widely used in life science and optical microscopy. However, di-sulfonated dyes were considered cell impermeable and not applicable in living cells. We challenged this assumption with 5 most popular rhodamines (Rho) having two carboxylic acid residues, versatile sulfonation patterns and emitting green (AS488), yellow (Rho530), orange (Rho565) and red (Rho590 and STAR RED) light. The probes comprising one rhodamine entity and a HaloTag^TM amine (O2) ligand (x) were prepared and applied for labeling of living, Vimentin-Halo (VIM-Halo) expressing U-2 OS cells. Surprisingly, we observed specific and bright staining with simplest compounds Rho590-x, Rho565-x and Rho530-x bearing two negative charges; they performed well also in stimulated emission depletion (STED) microscopy. Specific staining and red shifts in absorption and emission bands were observed with other probes having one negative charge; they were prepared by native chemical ligation and esterification.

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一个惊喜：基于二磺化若丹明和HaloTag的带负电荷荧光探针标记活细胞

二磺化若丹明是一种耐光且明亮的染料，广泛应用于生命科学和光学显微镜。然而，二磺化染料被认为是细胞不可渗透的，不适用于活细胞。我们用5种最流行的若丹明（Rho）挑战了这一假设，它们具有两个羧酸残基，多功能磺化模式，并发出绿色（AS488）、黄色（Rho530）、橙色（Rho565）和红色（Rho590和STAR red）光。制备包含一个罗丹明实体和HaloTag TM胺（O2）配体（x）的探针，并将其应用于表达活的Vimentin Halo（VIM Halo）的U-2 OS细胞的标记。令人惊讶的是，我们观察到带有两个负电荷的最简单的化合物Rho590-x、Rho565-x和Rho530-x的特异性和明亮的染色；它们在受激发射损耗（STED）显微镜中也表现良好。用具有一个负电荷的其他探针观察到吸收带和发射带的特异性染色和红移；它们是通过天然化学连接和酯化制备的。

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