ArticleNature methods2018
Precision and accuracy of single-molecule FRET measurements-a multi-laboratory benchmark study.
Erratum issuedOpen access · hybridAbstract read
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Article in Nature methods, 2018. The graph could read no effect estimate from its abstract, so it casts no vote on the map. An erratum has been issued. Cited by 260 papers.
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260citing papers in PubMed
57.3field-weighted citation impact, top 1% of its field
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Who cites it
260 citing papers in PubMed, 509 citations in OpenAlex.
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- DNA damage induces long range changes to duplex structure-a non-protein start to damage detection?Nucleic acids research · 2026Article
- Benchmarking dynamic analysis methods in diffusion-based single-molecule FRET.Cell reports methods · 2026Article
- Red-to-Near-Infrared Fluorescence Resonance Energy Transfer Reporters for Dual Imaging Applications.Small methods · 2026Article
- ATP-driven conformational dynamics reveal hidden intermediates in a heterodimeric ABC transporter.eLife · 2026Article
- Dynamic positioning of Rpc34 winged helix in RNA polymerase III elongation complex for its stability with implications for reinitiation.Proceedings of the National Academy of Sciences of the United States of America · 2026Article
- Single-molecule FRET and tracking of transfected biomolecules in living cells.Biophysical journal · 2026Article
- A Standardized Prism-Based TIRF Platform for Quantitative Single-Molecule Fluorescence Studies of Biomolecular Dynamics.Biosensors · 2026Article
- Dissecting mechanisms of ligand binding and conformational changes in the glutamine-binding protein.eLife · 2026Article
- Advancements in single-molecule fluorescence spectroscopy for probing conformations, dynamics, and interactions in disordered protein regions.Current opinion in structural biology · 2026Review
- A combined PIE-FRET and FCS assay to monitor RNA dynamics and cleavage by SARS-CoV-2 Nsp15.Scientific reports · 2026Article
- Cellular context and ligand class shape CXCR4-CCR5 heteromerization in live cells.Research square · 2026Article
- Ligand Binding Dynamics of Ion Channels and GPCRs Using Single-Molecule Fluorescence.Annual review of biophysics · 2026Review
- Resolving the ambiguous binding site of quercetin at the calcineurin subunit junction using funnel metadynamics with deep learning collective variables.Journal of computer-aided molecular design · 2026Article
- Single-Molecule Forster Resonance Energy Transfer With a Minimalistic 3D-Printed Setup and Dyes in the Blue-Green Spectral Region.Chemphyschem : a European journal of chemical physics and physical chemistry · 2026Article
- Article
- Unraveling allosteric signaling of G protein-coupled receptors (GPCRs) by single-molecule fluorescence.Biophysical reviews · 2026Review
- The Fantastic Single-Molecule Techniques.Chemical & biomedical imaging · 2026Review
- Kinesin-1 is highly flexible and adopts an open conformation in the absence of cargo.iScience · 2026Article
200 more citing papers are in PubMed but not listed here.
Corrections and comments
- Erratum issued
Authors and funding
63 authors at 20 institutions in 8 countries.
Björn HellenkampInstitute of Physical Chemistry, University of Freiburg, Freiburg im Breisgau, Germany.
Sonja SchmidInstitute of Physical Chemistry, University of Freiburg, Freiburg im Breisgau, Germany.ORCID http://orcid.org/0000-0002-3710-5602
Olga DoroshenkoMolecular Physical Chemistry, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, Germany.
Oleg OpanasyukMolecular Physical Chemistry, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, Germany.
Ralf KühnemuthMolecular Physical Chemistry, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, Germany.
Soheila Rezaei AdarianiDepartment of Physics and Astronomy, Clemson University, Clemson, SC, USA.
Benjamin AmbroseDepartment of Chemistry, University of Sheffield, Sheffield, UK.
Mikayel AznauryanInterdisciplinary Nanoscience Center (iNANO) and Department of Chemistry, Aarhus University, Aarhus, Denmark.ORCID http://orcid.org/0000-0002-5395-3441
Anders BarthPhysical Chemistry, Department of Chemistry, Nanosystems Initiative Munich (NIM), Center for Integrated Protein Science Munich (CiPSM) and Center for Nanoscience (CeNS), Ludwig-Maximilians-Universität München, Munich, Germany.ORCID http://orcid.org/0000-0003-3671-3072
Victoria BirkedalInterdisciplinary Nanoscience Center (iNANO) and Department of Chemistry, Aarhus University, Aarhus, Denmark.ORCID http://orcid.org/0000-0002-1360-8870
Mark E BowenDepartment of Physiology & Biophysics, Stony Brook University, Stony Brook, NY, USA.
Hongtao ChenDepartment of Biomedical Engineering, University of California, Irvine, Irvine, CA, USA.
Thorben CordesMolecular Microscopy Research Group, Zernike Institute for Advanced Materials, University of Groningen, Groningen, the Netherlands.
Tobias EilertInstitute for Biophysics, Ulm University, Ulm, Germany.
Carel FijenLaboratory of Biophysics, Wageningen University & Research, Wageningen, the Netherlands.
Christian GebhardtPhysical and Synthetic Biology, Faculty of Biology, Ludwig-Maximilians-Universität München, Planegg-Martinsried, Germany.
Markus GötzInstitute of Physical Chemistry, University of Freiburg, Freiburg im Breisgau, Germany.ORCID http://orcid.org/0000-0002-1956-7328
Giorgos GouridisMolecular Microscopy Research Group, Zernike Institute for Advanced Materials, University of Groningen, Groningen, the Netherlands.
Enrico GrattonDepartment of Biomedical Engineering, University of California, Irvine, Irvine, CA, USA.
Taekjip HaDepartment of Biomedical Engineering, John Hopkins University, Baltimore, MD, USA.ORCID http://orcid.org/0000-0003-2195-6258
Pengyu HaoDepartment of Physics, North Carolina State University, Raleigh, NC, USA.
Christian A HankeMolecular Physical Chemistry, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, Germany.
Andreas HartmannB CUBE-Center for Molecular Bioengineering, TU Dresden, Dresden, Germany.
Jelle HendrixLaboratory for Photochemistry and Spectroscopy, Department of Chemistry, University of Leuven, Leuven, Belgium.
Lasse L HildebrandtInterdisciplinary Nanoscience Center (iNANO) and Department of Chemistry, Aarhus University, Aarhus, Denmark.
Verena HirschfeldInstitute of Physics, University of Lübeck, Lübeck, Germany.
Johannes HohlbeinLaboratory of Biophysics, Wageningen University & Research, Wageningen, the Netherlands.
Boyang HuaDepartment of Biomedical Engineering, John Hopkins University, Baltimore, MD, USA.ORCID http://orcid.org/0000-0002-8869-9762
Christian G HübnerInstitute of Physics, University of Lübeck, Lübeck, Germany.
Eleni KallisInstitute for Biophysics, Ulm University, Ulm, Germany.
Achillefs N KapanidisGene Machines Group, Clarendon Laboratory, Department of Physics, University of Oxford, Oxford, UK.
Jae-Yeol KimSchool of Chemistry, Seoul National University, Seoul, South Korea.
Georg KrainerB CUBE-Center for Molecular Bioengineering, TU Dresden, Dresden, Germany.
Don C LambPhysical Chemistry, Department of Chemistry, Nanosystems Initiative Munich (NIM), Center for Integrated Protein Science Munich (CiPSM) and Center for Nanoscience (CeNS), Ludwig-Maximilians-Universität München, Munich, Germany.ORCID http://orcid.org/0000-0002-0232-1903
Nam Ki LeeSchool of Chemistry, Seoul National University, Seoul, South Korea.
Edward A LemkeDepartments of Biology and Chemistry, Pharmacy and Geosciences, Johannes Gutenberg-University Mainz, Mainz, Germany.ORCID http://orcid.org/0000-0002-0634-0503
Brié LevesqueDepartment of Physiology & Biophysics, Stony Brook University, Stony Brook, NY, USA.
Marcia LevitusSchool of Molecular Sciences and The Biodesign Institute, Arizona State University, Tempe, AZ, USA.
James J McCannDepartment of Physiology & Biophysics, Stony Brook University, Stony Brook, NY, USA.
Nikolaus Naredi-RainerPhysical Chemistry, Department of Chemistry, Nanosystems Initiative Munich (NIM), Center for Integrated Protein Science Munich (CiPSM) and Center for Nanoscience (CeNS), Ludwig-Maximilians-Universität München, Munich, Germany.
Daniel NettelsDepartment of Biochemistry, University of Zurich, Zurich, Switzerland.
Thuy NgoDepartment of Biomedical Engineering, John Hopkins University, Baltimore, MD, USA.
Ruoyi QiuDepartment of Physics, North Carolina State University, Raleigh, NC, USA.
Nicole C RobbGene Machines Group, Clarendon Laboratory, Department of Physics, University of Oxford, Oxford, UK.
Carlheinz RöckerInstitute for Biophysics, Ulm University, Ulm, Germany.
Hugo SanabriaDepartment of Physics and Astronomy, Clemson University, Clemson, SC, USA.ORCID http://orcid.org/0000-0001-7068-6827
Michael SchlierfB CUBE-Center for Molecular Bioengineering, TU Dresden, Dresden, Germany.ORCID http://orcid.org/0000-0002-6209-2364
Tim SchröderDepartment of Chemistry, Ludwig-Maximilians-Universität München, München, Germany.
Benjamin SchulerDepartment of Biochemistry, University of Zurich, Zurich, Switzerland.ORCID http://orcid.org/0000-0002-5970-4251
Henning SeidelInstitute of Physics, University of Lübeck, Lübeck, Germany.
Lisa StreitInstitute for Biophysics, Ulm University, Ulm, Germany.
Johann ThurnInstitute of Physical Chemistry, University of Freiburg, Freiburg im Breisgau, Germany.
Philip TinnefeldDepartment of Chemistry, Ludwig-Maximilians-Universität München, München, Germany.
Swati TyagiStructural and Computational Biology Unit, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
Niels VandenberkLaboratory for Photochemistry and Spectroscopy, Department of Chemistry, University of Leuven, Leuven, Belgium.
Andrés Manuel VeraDepartment of Chemistry, Ludwig-Maximilians-Universität München, München, Germany.
Keith R WeningerDepartment of Physics, North Carolina State University, Raleigh, NC, USA.
Bettina WünschInstitute of Physical & Theoretical Chemistry, Braunschweig Integrated Centre of Systems Biology (BRICS), and Laboratory for Emerging Nanometrology (LENA), Braunschweig University of Technology, Braunschweig, Germany.ORCID http://orcid.org/0000-0003-3654-4010
Inna S Yanez-OrozcoDepartment of Physics and Astronomy, Clemson University, Clemson, SC, USA.
Jens MichaelisInstitute for Biophysics, Ulm University, Ulm, Germany. jens.michaelis@uni-ulm.de.ORCID http://orcid.org/0000-0002-2739-4172
Claus A M SeidelMolecular Physical Chemistry, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, Germany. cseidel@hhu.de.ORCID http://orcid.org/0000-0002-5171-149X
Timothy D CraggsDepartment of Chemistry, University of Sheffield, Sheffield, UK. t.craggs@sheffield.ac.uk.ORCID http://orcid.org/0000-0002-7121-0609
Thorsten HugelInstitute of Physical Chemistry, University of Freiburg, Freiburg im Breisgau, Germany. thorsten.hugel@pc.uni-freiburg.de.ORCID http://orcid.org/0000-0003-3292-4569
Heinrich Heine University Düsseldorf · DEUniversität Ulm · DEUniversity of Freiburg · DELudwig-Maximilians-Universität München · DEAarhus University · DKCenter for Integrated Protein Science Munich · DEClemson University · USNorth Carolina State University · USStony Brook University · USUniversity of Lübeck · DEUniversity of Oxford · GBSeoul National University · KRTechnische Universität Dresden · DEUniversity of California, Irvine · USUniversity of Groningen · NLUniversity of Zurich · CHWageningen University & Research · NLArizona State University · USEuropean Molecular Biology Laboratory · DEHasselt University · BE
Funding
Tissue Structural and Neural Remodeling in Human Sacroiliac JointP20GM121342 · NIGMS · CLEMSON UNIVERSITY · PI Hai Yao · 2018 to 2026
$24.7MSINGLE MOLECULE ANALYSIS OF PSD-95 STRUCTURE AND LIGAND BINDINGR01MH081923 · NIMH · STATE UNIVERSITY NEW YORK STONY BROOK · PI BOWEN, MARK E · 2008 to 2022
$5.6MIntegrative single molecule studies: DNA repair and technology developmentR35GM127151 · NIGMS · UNIV OF NORTH CAROLINA CHAPEL HILL · PI DOROTHY A ERIE · 2018 to 2026
$4.6MSingle Molecule Studies of Nucleic Acids RemodelingR35GM122569 · NIGMS · JOHNS HOPKINS UNIVERSITY · PI Taekjip Ha · 2017 to 2026
$3.5MTrinucleotide Repeats: Atypical Structures, Diseases, and EvolutionR01GM118508 · NIGMS · NORTH CAROLINA STATE UNIVERSITY RALEIGH · PI SAGUI, MARIA CELESTE · 2016 to 2019
$1.1MQuantitative Imaging and Modeling of Regulation by Bacterial Small RNAR01GM112659 · NIGMS · UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN · PI HA, TAEKJIP, LUTHEY-SCHULTEN, ZAIDA · 2015 to 2018
$1.1MQuantitative analysis of transient DNA repair processes in vivoR01GM109832 · NIGMS · UNIV OF NORTH CAROLINA CHAPEL HILL · PI BRUSTAD, ERIC MICHAEL, ERIE, DOROTHY A · 2014 to 2017
$1.0MBiotechnology and Biological Sciences Research Council BB/H01795X/1European Research Council 261227European Research Council 681891NIGMS NIH HHS P20 GM121342NIGMS NIH HHS R01 GM109832NIGMS NIH HHS R01 GM112659NIGMS NIH HHS R01 GM118508NIGMS NIH HHS R35 GM122569NIGMS NIH HHS R35 GM127151NIMH NIH HHS R01 MH081923
Abstract
Single-molecule Förster resonance energy transfer (smFRET) is increasingly being used to determine distances, structures, and dynamics of biomolecules in vitro and in vivo. However, generalized protocols and FRET standards to ensure the reproducibility and accuracy of measurements of FRET efficiencies are currently lacking. Here we report the results of a comparative blind study in which 20 labs determined the FRET efficiencies (E) of several dye-labeled DNA duplexes. Using a unified, straightforward method, we obtained FRET efficiencies with s.d. between ±0.02 and ±0.05. We suggest experimental and computational procedures for converting FRET efficiencies into accurate distances, and discuss potential uncertainties in the experiment and the modeling. Our quantitative assessment of the reproducibility of intensity-based smFRET measurements and a unified correction procedure represents an important step toward the validation of distance networks, with the ultimate aim of achieving reliable structural models of biomolecular systems by smFRET-based hybrid methods.
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Fluorescence Resonance Energy TransferLaboratoriesReproducibility of Results
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