Fast, Bright, and Reversible Fluorescent Labeling of Rhodamine-Binding Proteins

Rhodamine dyes conjugated to targeting ligands can yield exceptionally bright fluorescent probes for live-cell imaging. However, the limited permeability of such rhodamine derivatives restricts their broader applications, particularly <i>in vivo</i>. Here, we present Rho-tag and SiR-tag, engineered protein tags derived from bacterial multidrug-resistant proteins that bind unsubstituted (silicon) rhodamines with nanomolar affinity. Unsubstituted (silicon) rhodamines readily cross membranes and enable rapid, reversible, and fluorogenic labeling of the tags in mammalian cells within seconds. The labeling of Rho-tag and SiR-tag is compatible with various super-resolution imaging methods and allows their use alongside self-labeling tags, such as HaloTag7 and SNAP-tag. The high affinity and specificity of both tags, combined with the permeability and outstanding spectroscopic properties of rhodamines, make them particularly attractive for <i>in vivo</i> bioimaging, as demonstrated by efficient fluorescent labeling in<i>C. elegans</i>embryos and zebrafish larvae.

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Publication Details

Journal
Journal of the American Chemical Society
Published
2025-12-30
DOI
https://doi.org/10.1021/jacs.5c18083
Citations
2
Primary Topic
Advanced Fluorescence Microscopy Techniques
Type
article
Field-Weighted Citation Impact
3.27
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article

Fast, Bright, and Reversible Fluorescent Labeling of Rhodamine-Binding Proteins

Claudia Catapano, Jonas Wilhelm, Marina S. Dietz, Mirosław Tarnawski et al.
2 citations
Journal of the American Chemical Society
Advanced Fluorescence Microscopy Techniques
3.27
article

Fast, Bright, and Reversible Fluorescent Labeling of Rhodamine-Binding Proteins

Claudia Catapano, Jonas Wilhelm, Marina S. Dietz, Mirosław Tarnawski, Silja Zedlitz, K. Johnsson, Carsten Hoege, L Dornfeld, Julien Hiblot, Julian Kompa, Nicola Porzberg, Lukas Merk, Simon Hans Lilje, David Jocher, Soohyen Jang, Mike Heilemann, Runyu Mao, Anthony A. Hyman
article en
2 citations

Abstract

Rhodamine dyes conjugated to targeting ligands can yield exceptionally bright fluorescent probes for live-cell imaging. However, the limited permeability of such rhodamine derivatives restricts their broader applications, particularly <i>in vivo</i>. Here, we present Rho-tag and SiR-tag, engineered protein tags derived from bacterial multidrug-resistant proteins that bind unsubstituted (silicon) rhodamines with nanomolar affinity. Unsubstituted (silicon) rhodamines readily cross membranes and enable rapid, reversible, and fluorogenic labeling of the tags in mammalian cells within seconds. The labeling of Rho-tag and SiR-tag is compatible with various super-resolution imaging methods and allows their use alongside self-labeling tags, such as HaloTag7 and SNAP-tag. The high affinity and specificity of both tags, combined with the permeability and outstanding spectroscopic properties of rhodamines, make them particularly attractive for <i>in vivo</i> bioimaging, as demonstrated by efficient fluorescent labeling in<i>C. elegans</i>embryos and zebrafish larvae.

Journal of the American Chemical SocietyVol. 148(1)
Goethe University Frankfurt (DE), Goethe-Institute United Kingdom (GB), Max Planck Institute for Medical Research (DE), Protein Express (United States) (US), Cellular Therapeutics (United Kingdom) (GB), Max Planck Institute of Molecular Cell Biology and Genetics (DE), Max Planck School Matter to Life, École Polytechnique Fédérale de Lausanne (CH)
Openalex Percentile: Top 6%
Advanced Fluorescence Microscopy Techniques
3.27
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