Chemical Physics 96 Springer Series in Chemical Physics

Springer Series in chemical physics 96 Springer Series in chemical physics

Series Editors: A. W. Castleman, Jr. J. P. Toennies K. Yamanouchi W. Zinth

The purpose of this series is to provide comprehensive up-to-date monographs in both well established disciplines and emerging research areas within the broad fields of chemical physics and physical chemistry. The books deal with both fun- damental science and applications, and may have either a theoretical or an experimental emphasis. They are aimed primarily at researchers and graduate students in chemical physics and related fields.

Please view available titles in Springer Series in Chemical Physics on series homepage http://www.springer.com/series/676 Astrid Graslund¨ Rudolf Rigler Jerker Widengren Editors

Single Molecule Spectroscopy in Chemistry, Physics and Biology

Nobel Symposium

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123 Editors ProfessorAstridGraslund¨ Professor Jerker Widengren Stockholm University Royal Institute or Technology (KTH) Department of Biophysics Department of Biomolecular Physics 10691 Stockholm, Sweden 10691 Stockholm, Sweden E-Mail: [email protected] E-Mail: [email protected] Professor Rudolf Rigler SwissFederalInstituteofTechnologyLausanne(EPFL) 1015 Lausanne, Switzerland E-Mail: [email protected]

Series Editors: Professor A.W. Castleman, Jr. Department of Chemistry, The Pennsylvania State University 152 Davey Laboratory, University Park, PA 16802, USA Professor J.P. Toennies Max-Planck-Institut fur¨ Stromungsforschung¨ Bunsenstrasse 10, 37073 Gottingen,¨ Germany Professor K. Yamanouchi University of Tokyo, Department of Chemistry Hongo 7-3-1, 113-0033 Tokyo, Japan ProfessorW.Zinth Universitat¨ Munchen,Institutf¨ ur¨ Medizinische Optik Ottingerstr.¨ 67, 80538 Munchen,¨ Germany

Springer Series in Chemical Physics ISSN 0172-6218 ISBN 978-3-642-02596-9 e-ISBN 978-3-642-02597-6 DOI 10.1007/978-3-642-02597-6 Springer Heidelberg Dordrecht London New York

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Springer is a part of Springer Science+Business Media (www.springer.com) Nobel Symposium, June 2008, at the S˚anga S¨aby Conference Foreword

By selecting the first week of June 2008 for the Nobel Symposium “Single Molecular Spectroscopy in Chemistry, Physics and Biology”, Rudolf Rigler, Jerker Widengren and Astrid Gräslund have once again won the top prize for Meeting Organizers, providing us with a Mediterranean climate on top of the warm hospitality that is unique to Sweden. The S˚anga Säby Conference Center was an ideal place to spend this wonderful week, and the comfort of this beautiful place blended perfectly with the high calibre of the scientific programme. It was a special privilege for me to be able to actively participate in this meeting on a field that is in many important ways complementary to my own research. I was impressed by the interdisciplinary ways in which single molecule spectroscopy has evolved and is currently pursued, with ingredients originating from physics, all branches of chemistry and a wide range of biological and biomedical research. A beautiful concert by Semmy Stahlhammer and Johan Ullén further extended the interdisciplinary character of the symposium. I would like to combine thanks to Rudolf, Jerker and Astrid with a glance into a future of other opportunities to enjoy top-level science combined with warm hospitality in the Swedish tradition.

Zürich, Kurt Wüthrich April 2009 Participants of the Nobel-Symposium 138: First row: Sarah Unterkofler, Anders Liljas, Xiao-Dong Su, Birgitta Rigler, Carlos Busta- mante, Toshio Yanagida, Steven Block, Xiaowei Zhuang, Sunney Xie. Second row: Ivan Scheblykin, Lars Thelander, Petra Schwille, Watt W. Webb, Rudolf Rigler, Jerker Widengren, Peter Lu, Shimon Weiss, William E Moerner, David Bensimon. Third row: Anders Ehrenberg, Yu Ming, Fredrik Elinder, Kazuhiko Kinosita, Vladana Vukojevic, Masataka Kinjo, May D Wang, Yu Ohsugi, Shuming Nie, Andreas Engel, Peter G Wolynes, Michel Orrit, Hans Blom, Johan Hofkens. Fourth row: Claus Seidel, Heike Hevekerl, Taekjip Ha, Evangelos Sisamakis, Per Ahlberg, Joseph Nordgren, Kurt Wthrich, Sune Svanberg, Bengt Nordn, Paul Alivisatos, Per Thyberg, Richard Keller, Andriy Chmyrov, Johan Elf, Per Rigler, Kai Hassler, Gustav Persson, Jürgen Köhler, Eric Betzig, Thomas Schmidt, Christoph Bräuchle, Elliot Elson, M˙ans Ehrenberg, Dimitrios K Papadopoulos, Ingemar Lundström, Horst Vogel, Stefan Wennmalm, Hermann Gaub, H˚akan Wennerström, Yosif Klafter, Julio Fernandez. Preface

The development of Single Molecule Detection and Spectroscopy started in the late eighties. The developments came from several areas. Fluorescence-based single molecule spectroscopy developed in particular from (i) holeburning and zero phonon spectroscopy of organic molecules at cryo temperatures and (ii) confocal fluctuation spectroscopy of emitting molecules at elevated temperatures. Of crucial importance for these approaches was the ability to suppress background radiation to the point where signals of single molecules could be detected. Today, confocal single molecule analysis is the dominating approach, particularly in chemistry and in biosciences, but attempts to combine analysis at low and high temperatures are being pursued. In parallel with this development, significant progress has been made in the field of single molecule force spectroscopy. Approaches based on atomic force microscopy, optical trapping, microneedles or magnetic beads have made it possible to investigate mechanical properties, and not least, the interplay between mechanics and chemistry on a single molecule level. In June 1999 the first Nobel Conference on Single Molecule Spectroscopy was organized in Södergarn Mansion, Lidingö (Sweden) and a comprehensive presentation of the results obtained in the first decade of single molecule analysis was given (Orrit, Rigler, Basche (eds.) 1999) Now after almost another decade, it was of interest to find out whether the developments and promises presented at the Södergarn Conference were still valid or had even exceeded our expectations. The contributions to this volume come from the pioneers of the early period of single molecule spectroscopy as well as from other laboratories which have made important contributions to demonstrate the importance of SM analysis in various applications in Chemistry, Physics and BioSciences. The Nobel Symposium No. 138 dedicated to Single Molecule Spectroscopy in Chemistry, Physics and Biosciences was held at the Mansion S˚anga-Säby situated at the island of EkeröinLakeMälaren outside Stockholm, from June 1–6, 2008. The Conference was blessed with pleasant weather and sunshine all the days. Together with the wonderful surroundings this contributed to many XPreface stimulating opportunities for individual discussions, in parallel with outdoor excursions including swimming in the lake, jogging tours, walks in the forests and sauna. The Symposium started with an evening session on molecules and dynamic processes by Kurt Wüthrich and Martin Karplus. The program of the next days included the presentation of the fields which initiated single molecule analysis in cryo temperatures (Moerner,Orrit) followed by confocal analysis of molecular fluctuations at room temperature (Keller, Rigler, Elson, Webb, Widengren, Schwille). Major topics in the following sessions included quantum dots (Alivasatos, Nie), the analysis of conformational dynamics (Weiss, Ha, Seidel), the motion of molecular motors (Yanagida, Kinosita) and repli- cating assemblies (Bustamante, Block). A special session was devoted to the analysis of forces operating on single molecules (Gaub, Fernandez) as well as to high resolution imaging of single molecules (Hell, Betzig, Zhuang, Engel). Stochastic single molecule events at the cellular level were another important topic (Xie, Schmidt, Vogel, Wolynes) as well as single molecule enzymology (Lu, Xie, Rigler, Hofkens, Klafter, Köhler), which together with atomic force microscopy formed the basis for intense discussions. Several presentations brought the single molecule methodologies and perspectives to a sub-cellular and cellular context (Rigler, Schwille, Weiss, Bensimon, Axner, Hell, Betzig, Zhuang, Schmidt, Xie, Orwar, Bräuchle), which seems to form one of several exciting future directions of this field. A special event was the evening concert with Semmy Stalhammer on the violin and Johan Ullén on the piano. The violin sonata of Cesar Franck and its masterly performance matched perfectly the level and tension of the scientific sessions. As organizers we would like to thank all the invited speakers for their excellent contributions to this symposium, as well as all those who contributed with a chapter to this book. We would also like to thank Margareta Klingberg and colleagues at the conference site of S˚anga-Säby for the prerequisites and support of an excellent venue, and not the least the Nobel Foundation for supporting this Symposium.

Stockholm, Astrid Gr¨aslund July 2009 Rudolf Rigler Jerker Widengren Contents

Part I Introductory Lecture: Molecular Dynamics of Single Molecules

1 How Biomolecular Motors Work: Synergy Between Single Molecule Experiments and Single Molecule Simulations Martin Karplus and Jingzhi Pu ...... 3

Part II Detection of Single Molecules and Single Molecule Processes

2 Single-Molecule Optical Spectroscopy and Imaging: From Early Steps to Recent Advances William E. Moerner ...... 25 3 Single Molecules as Optical Probes for Structure and Dynamics Michel Orrit...... 61 4 FCS and Single Molecule Spectroscopy Rudolf Rigler ...... 77

Part III Fluorescence-Correlation Spectroscopy

5 Single-Molecule Spectroscopy Illuminating the Molecular Dynamics of Life Watt W. Webb ...... 107 6 Chemical Fluxes in Cellular Steady States Measured by Fluorescence-Correlation Spectroscopy Hong Qian and Elliot L. Elson ...... 119 XII Contents

7 In Vivo Fluorescence Correlation and Cross-Correlation Spectroscopy Jörg Mütze, Thomas Ohrt, Zdenˇek Petráˇsek, and Petra Schwille ...... 139 8 Fluorescence Flicker as a Read-Out in FCS: Principles, Applications, and Further Developments Jerker Widengren ...... 155

Part IV Quantum Dots and Single Molecule Behaviour

9 Development of Nanocrystal Molecules for Plasmon Rulers and Single Molecule Biological Imaging A.P. Alivisatos ...... 175 10 Size-Minimized Quantum Dots for Molecular and Cellular Imaging Andrew M. Smith, Mary M. Wen, May D. Wang, and Shuming Nie ....187 11 Mapping Transcription Factors on Extended DNA: A Single Molecule Approach Yuval Ebenstein, Natalie Gassman, and Shimon Weiss ...... 203

Part V Molecular Motion of Contractile Elements and Polymer Formation

12 Single-Molecule Measurement, a Tool for Exploring the Dynamic Mechanism of Biomolecules Toshio Yanagida ...... 219 13 Viral DNA Packaging: One Step at a Time Carlos Bustamante and Jeﬀrey R. Moﬃtt ...... 237 14 Chemo-Mechanical Coupling in the Rotary Molecular Motor F1-ATPase Kengo Adachi, Shou Furuike, Mohammad Delawar Hossain, Hiroyasu Itoh, Kazuhiko Kinosita, Jr., Yasuhiro Onoue, and Rieko Shimo-Kon ...271

Part VI Force and Multiparameter Spectroscopy on Functional Active Proteins

15 Mechanoenzymatics and Nanoassembly of Single Molecules Elias M. Puchner and Hermann E. Gaub ...... 289 Contents XIII

16 Single Cell Physiology Pierre Neveu, Deepak Kumar Sinha, Petronella Kettunen, Sophie Vriz, Ludovic Jullien, and David Bensimon ...... 305 17 Force-Clamp Spectroscopy of Single Proteins Julio M Fernandez, Sergi Garcia-Manyes, and Lorna Dougan ...... 317 18 Unraveling the Secrets of Bacterial Adhesion Organelles Using Single-Molecule Force Spectroscopy Ove Axner, Oscar Björnham, Mickaël Castelain, Efstratios Koutris, Staffan Schedin, Erik Fällman, and Magnus Andersson ...... 337

Part VII Nanoscale Microscopy and High Resolution Imaging

19 Far-Field Optical Nanoscopy Stefan W. Hell ...... 365 20 Sub-Diﬀraction-Limit Imaging with Stochastic Optical Reconstruction Microscopy Mark Bates, Bo Huang, Michael J. Rust, Graham T. Dempsey, Wenqin Wang, and Xiaowei Zhuang ...... 399 21 Assessing Biological Samples with Scanning Probes A. Engel ...... 417

Part VIII Single Molecule Microscopy in Individual Cells

22 Enzymology and Life at the Single Molecule Level X. Sunney Xie ...... 435 23 Controlling Chemistry in Dynamic Nanoscale Systems Aldo Jesorka, Ludvig Lizana, Zoran Konkoli, Ilja Czolkos, and Owe Orwar ...... 449

Part IX Catalysis of Single Enzyme Molecules

24 Single-Molecule Protein Conformational Dynamics in Enzymatic Reactions H. Peter Lu ...... 471 25 Watching Individual Enzymes at Work Kerstin Blank, Susana Rocha, Gert De Cremer, Maarten B.J. Roeﬀaers, Hiroshi Uji-i, and Johan Hofkens ...... 495 XIV Contents

26 The Influence of Symmetry on the Electronic Structure of the Photosynthetic Pigment-Protein Complexes from Purple Bacteria Martin F. Richter, Jürgen Baier, Richard J. Cogdell, Silke Oellerich, and Jürgen Köhler ...... 513

Part X Fields and Outlook

27 Exploring Nanostructured Systems with Single-Molecule Probes: From Nanoporous Materials to Living Cells Christoph Br¨auchle ...... 537 28 Gene Regulation: Single-Molecule Chemical Physics in a Natural Context Peter G. Wolynes ...... 553

Index ...... 561 Contributors

Kengo Adachi Universität Bayreuth Department of Physics Universtitätsstrasse 30 Faculty of Science and Engineering 95440 Bayreuth, Germany Waseda University, Okubo Shinjuku-ku, Tokyo 169-8555 Mark Bates Japan School of Engineering and Applied A. P. Alivisatos Sciences Department of Chemistry 29 Oxford Street, Cambridge University of California MA 02138, USA Berkeley, USA and David Bensimon Materials Science Division Laboratoire de Physique Statistique Lawrence Berkeley National Lab UMR 8550 Berkeley, USA Ecole Normale Supérieure [email protected] Paris, France and Magnus Andersson Department of Physics Department of Chemistry and Ume˚aUniversity Biochemistry 901 87 Ume˚a, Sweden University of California at Los Angeles Ove Axner Los Angeles, CA, USA Department of Physics [email protected], Ume˚aUniversity [email protected] 901 87 Ume˚a, Sweden [email protected] Oscar Björnham Jürgen Baier Department of Applied Physics Experimental Physics IV and Electronics and Bayreuth Institute for Ume˚aUniversity Macromolecular Research 901 87 Ume˚a, Sweden XVI Contributors

Kerstin Blank Gert De Cremer Department of Chemistry Department of Microbial and Katholieke Universiteit Leuven Molecular Systems Leuven, Belgium Katholieke Universiteit Leuven Leuven, Belgium Christoph Bräuchle Ilja Czolkos Department of Chemistry und Department of Physical Chemistry Biochemistry and Center for Chalmers University of Technology Nanoscience (CeNS) 412 96 Gothenburg, Sweden Ludwig-Maximilians-Universität München Graham T. Dempsey Butenandtstrasse 11 Program in Biophysics 81377 München, Germany Harvard University, Cambridge Christoph.Braeuchle@ MA 02138, USA cup.uni-muenchen.de Lorna Dougan Department of Biological Sciences Carlos Bustamante Columbia University Jason L. Choy Laboratory of Single New York, NY 10027, USA Molecule Biophysics and Department Yuval Eb enstein of Physics Department of Chemistry and University of California, Berkeley Biochemistry CA 94720, USA and DOE Institute for Genomics and and Proteomics Departments of Chemistry and UCLA, Germany Molecular and Cell Biology Elliot L. Elson Howard Hughes Medical Institute Department of Biochemistry and University of California, Berkeley Molecular Biophysics CA 94720, USA Washington University [email protected] St. Louis, MO 63110, USA [email protected] Mickaël Castelain Department of Physics A. Engel Ume˚aUniversity Maurice E. Müller Institute for 901 87 Ume˚a, Sweden Structural Biology Biozentrum, University of Basel Klingelbergstrasse 70, 4056 Basel Richard J. Cogdell Switzerland Division of Biochemistry and Molecular Biology and Institute of Biomedical and Life Department of Pharmacology Sciences Case Western Reserve University Biomedical Research Building 10900 Euclid Avenue University of Glasgow Wood Bldg 321D, Cleveland 120 University Place OH 44106, USA Glasgow G12 8TA, UK [email protected] Contributors XVII

Erik Fällman Johan Hofkens Department of Physics Department of Chemistry Ume˚aUniversity Katholieke Universiteit Leuven 901 87 Ume˚a, Sweden Leuven, Belgium Julio M Fernandez [email protected]. Department of Biological Sciences ac.be Columbia University, New York NY 10027, USA Mohammad Delawar Hossain [email protected] Department of Physics Shou Furuike Faculty of Science and Engineering Department of Physics Waseda University, Okubo Faculty of Science and Engineering Shinjuku-ku, Tokyo 169-8555 Waseda University, Okubo Japan Shinjuku-ku, Tokyo 169-8555 and Japan Department of Physics Sergi Garcia-Manyes School of Physical Sciences Department of Biological Sciences Shahjalal University of Science and Columbia University Technology New York, NY 10027, USA Sylhet-3114, Bangladesh Natalie Gassman Department of Chemistry and Bo Huang Biochemistry Department of Chemistry and and DOE Institute for Genomics and Chemical Biology Proteomics Cornell University, Ithaca UCLA, Germany NY, USA Hermann E. Gaub and Lehrstuhl für Angewandte Physik Howard Hughes Medical Institute LMU Munich, Amalienstr. 54 Harvard University, Cambridge 80799 Munich, Germany MA 02138, USA and [email protected] Center for Nanoscience (CENS) Nanosystems Initiative Munich (NIM) and Center for Integrated Hiroyasu Itoh Protein Science Munich (CIPSM) Tsukuba Research Laboratory Germany Hamamatsu Photonics KK [email protected] Tokodai, Tsukuba 300-2635 Japan Stefan W. Hell Department of NanoBiophotonics Max Planck Institute for Biophysical Aldo Jesorka Chemistry Department of Physical Chemistry 37070 Göttingen, Germany Chalmers University of Technology [email protected],[email protected] 412 96 Gothenburg, Sweden XVIII Contributors

Ludovic Jullien Efstratios Koutris Département de Chimie UMR 8640 Department of Physics Ecole Normale Supérieure Ume˚aUniversity Paris, France 901 87 Ume˚a, Sweden [email protected] Ludvig Lizana Martin Karplus Department of Physical Chemistry Department of Chemistry and Chalmers University of Technology Chemical Biology 412 96 Gothenburg, Sweden Harvard University, Cambridge MA 02138, USA [email protected] H. Peter Lu Department of Chemistry Petronella Kettunen Center for Photochemical Sciences Department of Physiological Science Bowling Green State University University of California at Los Bowling Green Angeles OH 43403, USA Los Angeles, CA, USA [email protected] [email protected] William E. Moerner Kazuhiko Kinosita, Jr Departments of Chemistry and Department of Physics (by Courtesy) of Applied Physics Faculty of Science and Engineering Stanford University, Stanford Waseda University, Okubo CA 94305, USA Shinjuku-ku, Tokyo 169-8555 [email protected] Japan [email protected] http://www.k2.phys.waseda.ac.jp Jeffrey R. Moffitt Jason L. Choy Laboratory of Single Molecule Biophysics and Department Jürgen Köhler of Physics Experimental Physics IV University of California and Bayreuth Institute for Berkeley Macromolecular Research CA 94720, USA Universität Bayreuth Universtitätsstrasse 30 95440 Bayreuth, Germany Jörg Mütze [email protected] Biophysics group Biotechnologisches Zentrum Zoran Konkoli Technische Universität Dresden Microtechnology and Nanoscience Tatzberg 47-51 Center 01307 Dresden Chalmers University of Technology Germany, 412 96 Gothenburg petra.schwille@ Sweden biotec.tu-dresden.de Contributors XIX

Pierre Neveu Department of Functional Molecular Kavli Institute for Theoretical Science Physics The Graduate University for University of California at Santa Advanced Studies (Sokendai) Barbara Okazaki, Aichi 444-8585 Santa Barbara Japan CA, USA [email protected] Michel Orrit MoNOS, LION Shuming Nie Postbox 9504, Leiden University Departments of Biomedical 2300 RA Leiden Engineering and Chemistry The Netherlands Emory University and Georgia [email protected] Institute of Technology 101 Woodruff Circle Owe Orwar Suite 2001, Atlanta Department of Physical Chemistry GA 30322, USA Chalmers University of Technology [email protected] 412 96 Gothenburg, Sweden [email protected] Silke Oellerich Experimental Physics IV Zdenˇek Petráˇsek and Bayreuth Institute for Biophysics group Macromolecular Research Biotechnologisches Zentrum Universität Bayreuth Technische Universität Dresden Universtitätsstrasse 30 Tatzberg 47-51 95440 Bayreuth 01307 Dresden, Germany Germany petra.schwille@ biotec.tu-dresden.de Thomas Ohrt Biophysics group Jingzhi Pu Biotechnologisches Zentrum Laboratoire de Chimie Biophysique Technische Universität Dresden ISIS, UniversitéLouisPasteur Tatzberg 47-51, 01307 Dresden 67000 Strasbourg, France Germany, petra.schwille@ Elias M. Puchner biotec.tu-dresden.de Lehrstuhl für Angewandte Physik LMU Munich, Amalienstr. 54 Yasuhiro Onoue 80799 Munich, Germany Department of Physics and Faculty of Science and Engineering Center for Nanoscience (CENS) Waseda University, Okubo Nanosystems Initiative Munich Shinjuku-ku, Tokyo 169-8555 (NIM) and Center for Integrated Japan Protein Science Munich (CIPSM) and Germany XX Contributors

Hong Qian Rieko Shimo-Kon Department of Applied Mathematics Department of Physics University of Washington Faculty of Science and Engineering Seattle, WA 98195, USA Waseda University Okubo, Shinjuku-ku Martin F. Richter Tokyo 169-8555, Japan Experimental Physics IV and Bayreuth Institute for Deepak Kumar Sinha Macromolecular Research Laboratoire de Physique Statistique Universität Bayreuth UMR 8550 Universtitätsstrasse 30 Ecole Normale Supérieure 95440 Bayreuth, Germany Paris, France [email protected] Susana Rocha Andrew M. Smith Department of Chemistry Departments of Biomedical Katholieke Universiteit Leuven Engineering and Chemistry Leuven, Belgium Emory University and Georgia Institute of Technology Maarten B. J. Roeffaers 101 Woodruff Circle Department of Chemistry Suite 2001, Atlanta Katholieke Universiteit Leuven GA 30322, USA Leuven, Belgium Hiroshi Uji-i Michael J. Rust Department of Chemistry Department of Physics Harvard Katholieke Universiteit Leuven University Leuven, Belgium Cambridge, MA 02138 USA Sophie Vriz Inserm U770 Staffan Schedin Hémostase et Dynamique Cellulaire Department of Applied Physics and Vasculaire Electronics Le Kremlin-Bicêtre Ume˚aUniversity France 901 87 Ume˚a, Sweden [email protected]

May D. Wang Petra Schwille Departments of Biomedical Biophysics group Engineering Biotechnologisches Zentrum Georgia Institute of Technology Technische Universit¨at Dresden 313 Ferst Drive Tatzberg 47-51, 01307 Dresden UA Whitaker Building 4106 Germany Atlanta, GA 30332, USA petra.schwille@ biotec.tu-dresden.de and Contributors XXI

Department of Electrical and Peter G. Wolynes Computer Engineering Department of Chemistry and Georgia Institute of Technology Biochemistry 313 Ferst Drive University of California at San Diego UA Whitaker Building 4106 9500 Gilman Drive Atlanta, GA 30332, USA La Jolla, CA 92093 USA Wenqin Wang [email protected] Department of Physics X. Sunney Xie Harvard University Department of Chemistry and Cambridge Chemical Biology MA 02138, USA Harvard University Watt W. Webb Cambridge Cornell University MA 02138 USA School of Applied and Engineering [email protected] Physics Toshio Yanagida 212 Clark Hall Graduate School of Frontier Ithaca, NY 14853-2501, USA Biosciences [email protected] Osaka University, 1-3 Yamadaoka, Suita, Osaka Shimon Weiss 565-0871 Japan Department of Chemistry and and Biochemistry and DOE Institute for Genomics and Formation of soft nano-machines Proteomics CREST 1-3 Yamadaoka UCLA, University of California Suita, Osaka Los Angeles, CA, USA 565-0871 Japan [email protected] [email protected]. osaka-u.ac.jp Mary M. Wen http://www.phys1.med. Departments of Biomedical osaka-u.ac.jp/ Engineering and Chemistry Xiaowei Zhuang Emory University and Georgia Department of Chemistry and Institute of Technology Chemical Biology 101 Woodruﬀ Circle Howard Hughes Medical Institute Suite 2001, Atlanta Harvard University GA 30322, USA Cambridge, MA 02138 USA Jerker Widengren Exp.Biomol.Physics Dept. Appl. and Physics Department of Physics Royal Institute of Technology (KTH) Program in Biophysics Albanova University Center Harvard University, Cambridge, 106 91 Stockholm, Sweden MA 02138, USA [email protected] [email protected]