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      SiR–Hoechst is a far-red DNA stain for live-cell nanoscopy

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          Abstract

          Cell-permeable DNA stains are popular markers in live-cell imaging. Currently used DNA stains for live-cell imaging are either toxic, require illumination with blue light or are not compatible with super-resolution microscopy, thereby limiting their utility. Here we describe a far-red DNA stain, SiR–Hoechst, which displays minimal toxicity, is applicable in different cell types and tissues, and is compatible with super-resolution microscopy. The combination of these properties makes this probe a powerful tool for live-cell imaging.

          Abstract

          Existing DNA stains for live cell microscopy are either toxic, require illumination with blue light, or are not compatible with super-resolution microscopy. Here the authors develop SiRHoechst, a non-toxic far-red DNA stain that is compatible with super-resolution microscopy.

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          The fluorescent toolbox for assessing protein location and function.

          Ben Giepmans, Stephen R Adams, Mark Ellisman … (2006)
          Advances in molecular biology, organic chemistry, and materials science have recently created several new classes of fluorescent probes for imaging in cell biology. Here we review the characteristic benefits and limitations of fluorescent probes to study proteins. The focus is on protein detection in live versus fixed cells: determination of protein expression, localization, activity state, and the possibility for combination of fluorescent light microscopy with electron microscopy. Small organic fluorescent dyes, nanocrystals ("quantum dots"), autofluorescent proteins, small genetic encoded tags that can be complexed with fluorochromes, and combinations of these probes are highlighted.
          Article 0 comments Cited 492 times – based on 0 reviews     Review now
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            A general method to improve fluorophores for live-cell and single-molecule microscopy

            Jonathan Grimm, Brian English, Jiji Chen … (2015)
            Specific labeling of biomolecules with bright fluorophores is the keystone of fluorescence microscopy. Genetically encoded self-labeling tag proteins can be coupled to synthetic dyes inside living cells, resulting in brighter reporters than fluorescent proteins. Intracellular labeling using these techniques requires cell-permeable fluorescent ligands, however, limiting utility to a small number of classic fluorophores. Here, we describe a simple structural modification that improves the brightness and photostability of dyes while preserving spectral properties and cell permeability. Inspired by molecular modeling, we replaced the N,N-dimethylamino substituents in tetramethylrhodamine with four-membered azetidine rings. This addition of two carbon atoms doubles the quantum efficiency and improves the photon yield of the dye in applications ranging from in vitro single-molecule measurements to super-resolution imaging. The novel substitution is generalizable, yielding a palette of chemical dyes with improved quantum efficiencies that spans the UV and visible range.
            Article 0 comments Cited 461 times – based on 0 reviews     Review now
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              Fluorogenic probes for live-cell imaging of the cytoskeleton.

              Gražvydas Lukinavičius, Luc Reymond, Elisa D'Este … (2014)
              We introduce far-red, fluorogenic probes that combine minimal cytotoxicity with excellent brightness and photostability for fluorescence imaging of actin and tubulin in living cells. Applied in stimulated emission depletion (STED) microscopy, they reveal the ninefold symmetry of the centrosome and the spatial organization of actin in the axon of cultured rat neurons with a resolution unprecedented for imaging cytoskeletal structures in living cells.
              Article 0 comments Cited 302 times – based on 0 reviews     Review now
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                Author and article information

                Journal
                Journal ID (nlm-ta): Nat Commun
                Journal ID (iso-abbrev): Nat Commun
                Title: Nature Communications
                Publisher: Nature Pub. Group
                ISSN (Electronic): 2041-1723
                Publication date (Electronic): 01 October 2015
                Publication date Collection: 2015
                Volume: 6
                Electronic Location Identifier: 8497
                Affiliations
                [1 ]Ecole Polytechnique Fédérale de Lausanne (EPFL), Institute of Chemical Sciences and Engineering (ISIC), Institute of Bioengineering, NCCR in Chemical Biology , 1015 Lausanne, Switzerland
                [2 ]Institute of Molecular Biotechnology of the Austrian Academy of Sciences (IMBA), Vienna Biocenter Campus (VBC) , Dr Bohr Gasse 3, 1030 Vienna, Austria
                [3 ]Department of Chemistry—BMC, Uppsala University , 75123 Uppsala, Sweden
                [4 ]Department of Biochemistry, NCCR in Chemical Biology, University of Geneva , 1211 Geneva, Switzerland
                [5 ]Department NanoBiophotonics, Max-Planck-Institute for Biophysical Chemistry , Am Fassberg 11, 37077 Göttingen, Germany
                Author notes
                Article
                Publisher Item ID: ncomms9497
                DOI: 10.1038/ncomms9497
                PMC ID: 4600740
                PubMed ID: 26423723
                SO-VID: 690a4b85-ec32-4778-b213-c95a4eaf460d
                Copyright © Copyright © 2015, Nature Publishing Group, a division of Macmillan Publishers Limited. All Rights Reserved.
                License:

                This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article's Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/

                History
                Date received : 15 July 2015
                Date accepted : 27 August 2015
                Categories
                Subject: Article

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