Download or read book Single molecule and Super resolution in Living Cells written by Hsiao-lu Lee and published by . This book was released on 2011 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt: Since the first successful detection single molecules over two decades ago, single-molecule spectroscopy has developed into a burgeoning field with a wealth of experiments at room temperature and inside living cells. Probing asynchronous and heterogeneous populations in situ, one molecule at a time, is not only desirable, but critical for many biological questions. Further, super-resolution imaging based on sequential imaging of sparse subsets of single molecules, has seen explosive growth within the last five years. This dissertation describes both the application of live-cell single-molecule imaging as an answer to important biological questions, and development and validation of fluorescent probes for targeted super-resolution imaging.
Download or read book Single molecule and Super resolution Microscopy of Bacterial Cells written by Marissa Kim Lee and published by . This book was released on 2015 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt: Single molecules were first detected at low temperatures twenty-six years ago in the laboratory of W.E. Moerner. Subsequent technological advances have allowed researchers to study single molecules at room temperatures and within living cells, providing novel biological insight about underlying spatial and dynamical heterogeneity. By combining single molecule detection with the ability to control the emissive state of the fluorescent label (also called "active control"), a suite of super-resolution imaging techniques has been developed. These single-molecule-based super-resolution imaging strategies leverage the fluorescence microscope's ability to non-invasively study multiple targets within living cells, while bridging the resolution gap between optical and electron microscopies. In large part, future advances to improve single molecule and super-resolution imaging require better fluorophore and labeling technologies. Utilizing fluorophore with higher photon yields will increase the resolution of super-resolution images and the data acquisition speed. Additionally, a greater library fluorophores with different of colors and sensing capabilities will enable application to more imaging targets. Currently, many single molecule and super-resolution experiments within living systems use fluorescent proteins because the labeling of target proteins is more straightforward. However, the limited photon yield of fluorescent proteins often results in tantalizingly fuzzy super-resolution images. Imaging the same targets, labeled instead with brighter organic emitters, could provide more image detail, but better fluorogenic and genetically encoded labeling schemes must be developed and discovered. The first chapter of this dissertation will introduce and discuss the historical context and basic principles of single molecule and super-resolution imaging. Chapter 2 will then describe the general experimental procedures necessary for quantitative single molecule and super-resolution imaging, including quantifying the number of photons detected (and emitted) from a single molecule, as well as the preparation of bacterial samples for fluorescence microscopy. Later chapters apply these fundamental experimental measurements to study bacterial biology and fluorophore photophysics. Chapters 3 and 4 concern the development and characterization of organic emitters suitable for single molecule or super-resolution imaging, work achieved with the synthetic collaboration of organic chemists in the laboratory of Professor Robert J. Twieg at Kent State University. Chapter 3 discusses the optimization of rhodamine spirolactam photoswitching such that activation could occur at visible wavelengths. The optimized rhodamine spirolactams were then covalently attached to the surface of bacterial cells and imaged with three-dimensional super-resolution. Images of the bacterial cell surface demonstrates a marked improvement in labeling uniformity, specificity, and density compared to previous methods which labeled the surface with the transient binding of a membrane sensitive dye. Chapter 4 introduces a novel enzyme-based strategy to control the fluorescence from nitro-aryl fluorogens. A proof-of-principle experiment demonstrated that endogenous nitroreductase enzymes within bacterial cells could catalyze the fluorescence-activating reaction, thus generating free fluorophores, which were detectable on the single-molecule-level within the cell. Lastly, chapter 5 summarizes three-dimensional imaging experiments (performed in collaboration with the laboratory of Professor Lucy Shapiro in the Department of Developmental Biology at Stanford University) of components of the bacterial gene expression machinery labeled with fluorescent proteins. Super-resolution imaging is ideally suited to the small size scale of bacterial cells, and a wealth of biological insights remains to be discovered. Simultaneously improving fluorophore photon yield, specificity, and active control strategies will have a profound impact on super-resolution precision and speed.
Download or read book Single molecule Techniques written by Paul R. Selvin and published by CSHL Press. This book was released on 2008 with total page 511 pages. Available in PDF, EPUB and Kindle. Book excerpt: Geared towards research scientists in structural and molecular biology, biochemistry, and biophysics, this manual will be useful to all who are interested in observing, manipulating and elucidating the molecular mechanisms and discrete properties of macromolecules.
Download or read book Single Molecule Science written by Krishnarao Appasani and published by Cambridge University Press. This book was released on 2022-05-26 with total page 171 pages. Available in PDF, EPUB and Kindle. Book excerpt: A comprehensive volume that brings together authoritative overviews of single molecule science techniques from a biological perspective.
Download or read book Bioconjugate Techniques written by Greg T. Hermanson and published by Academic Press. This book was released on 2010-07-26 with total page 1233 pages. Available in PDF, EPUB and Kindle. Book excerpt: Bioconjugate Techniques, 2nd Edition, is the essential guide to the modification and cross linking of biomolecules for use in research, diagnostics, and therapeutics. It provides highly detailed information on the chemistry, reagent systems, and practical applications for creating labeled or conjugate molecules. It also describes dozens of reactions with details on hundreds of commercially available reagents and the use of these reagents for modifying or cross linking peptides and proteins, sugars and polysaccharides, nucleic acids and oligonucleotides, lipids, and synthetic polymers. A one-stop source for proven methods and protocols for synthesizing bioconjugates in the lab Step-by-step presentation makes the book an ideal source for researchers who are less familiar with the synthesis of bioconjugates More than 600 figures that visually describe the complex reactions associated with the synthesis of bioconjugates Includes entirely new chapters on the latest areas in the field of bioconjugation as follows: Microparticles and nanoparticlesSilane coupling agentsDendrimers and dendronsChemoselective ligationQuantum dotsLanthanide chelatesCyanine dyesDiscrete PEG compoundsBuckyballs,fullerenes, and carbon nanotubesMass tags and isotope tagsBioconjugation in the study of protein interactions
Download or read book Handbook of Single Molecule Biophysics written by Peter Hinterdorfer and published by Springer Science & Business Media. This book was released on 2009-12-24 with total page 634 pages. Available in PDF, EPUB and Kindle. Book excerpt: This handbook describes experimental techniques to monitor and manipulate individual biomolecules, including fluorescence detection, atomic force microscopy, and optical and magnetic trapping. It includes single-molecule studies of physical properties of biomolecules such as folding, polymer physics of protein and DNA, enzymology and biochemistry, single molecules in the membrane, and single-molecule techniques in living cells.
Download or read book Spectroscopy and Dynamics of Single Molecules written by and published by Elsevier. This book was released on 2019-08-14 with total page 404 pages. Available in PDF, EPUB and Kindle. Book excerpt: Spectroscopy and Dynamics of Single Molecules: Methods and Applications reviews the most recent developments in spectroscopic methods and applications. Spectroscopic techniques are the chief experimental methods for testing theoretical models and research in this area plays an important role in stimulating new theoretical developments in physical chemistry. This book provides an authoritative insight into the latest advances in the field, highlighting new techniques, current applications, and potential future developments An ideal reference for chemists and physicists alike, Spectroscopy and Dynamics of Single Molecules: Methods and Applications is a useful guide for all those working in the research, design, or application of spectroscopic tools and techniques across a wide range of fields. - Includes the latest research on ultrafast vibrational and electronic dynamics, nonlinear spectroscopies, and single-molecule methods - Makes the content accessible to researchers in chemistry, biophysics, and chemical physics through a rigorous multi-disciplinary approach - Provides content edited by a world-renowned chemist with more than 30 years of experience in research and instruction
Download or read book Single Molecule Tools Part B Super Resolution Particle Tracking Multiparameter and Force Based Methods written by and published by Academic Press. This book was released on 2010-07-09 with total page 745 pages. Available in PDF, EPUB and Kindle. Book excerpt: Single molecule tools have begun to revolutionize the molecular sciences, from biophysics to chemistry to cell biology. They hold the promise to be able to directly observe previously unseen molecular heterogeneities, quantitatively dissect complex reaction kinetics, ultimately miniaturize enzyme assays, image components of spatially distributed samples, probe the mechanical properties of single molecules in their native environment, and "just look at the thing" as anticipated by the visionary Richard Feynman already half a century ago. Single Molecule Tools, Part B: Super-Resolution, Particle Tracking, Multiparameter, and Force Based Methods captures a snapshot of this vibrant, rapidly expanding field, presenting articles from pioneers in the field intended to guide both the newcomer and the expert through the intricacies of getting single molecule tools. - Includes time-tested core methods and new innovations applicable to any researcher employing single molecule tools - Methods included are useful to both established researchers and newcomers to the field - Relevant background and reference information given for procedures can be used as a guide to developing protocols in a number of disciplines
Download or read book Far Field Optical Nanoscopy written by Philip Tinnefeld and published by Springer. This book was released on 2015-02-07 with total page 340 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book describes developments in the field of super-resolution fluorescence microscopy or nanoscopy. In 11 chapters, distinguished scientists and leaders in their respective fields describe different nanoscopy approaches, various labeling technologies, and concrete applications. The topics covered include the principles and applications of the most popular nanoscopy techniques STED and (f)PALM/STORM, along with advances brought about by fluorescent proteins and organic dyes optimized for fluorescence nanoscopy. Furthermore, the photophysics of fluorescent labels is addressed, specifically for improving their photoswitching capabilities. Important applications are also discussed, such as the tracking and counting of molecules to determine acting forces in cells, and quantitative cellular imaging, respectively, as well as the mapping of chemical reaction centers at the nano-scale. The 2014 Chemistry Nobel Prize® was awarded for the ground-breaking developments of super-resolved fluorescence microscopy. In this book, which was co-edited by one of the prize winners, readers will find the most recent developments in this field.
Download or read book Nanoscale Photonic Imaging written by Tim Salditt and published by Springer Nature. This book was released on 2020-06-09 with total page 634 pages. Available in PDF, EPUB and Kindle. Book excerpt: This open access book, edited and authored by a team of world-leading researchers, provides a broad overview of advanced photonic methods for nanoscale visualization, as well as describing a range of fascinating in-depth studies. Introductory chapters cover the most relevant physics and basic methods that young researchers need to master in order to work effectively in the field of nanoscale photonic imaging, from physical first principles, to instrumentation, to mathematical foundations of imaging and data analysis. Subsequent chapters demonstrate how these cutting edge methods are applied to a variety of systems, including complex fluids and biomolecular systems, for visualizing their structure and dynamics, in space and on timescales extending over many orders of magnitude down to the femtosecond range. Progress in nanoscale photonic imaging in Göttingen has been the sum total of more than a decade of work by a wide range of scientists and mathematicians across disciplines, working together in a vibrant collaboration of a kind rarely matched. This volume presents the highlights of their research achievements and serves as a record of the unique and remarkable constellation of contributors, as well as looking ahead at the future prospects in this field. It will serve not only as a useful reference for experienced researchers but also as a valuable point of entry for newcomers.
Download or read book Super Resolution Imaging in Biomedicine written by Alberto Diaspro and published by CRC Press. This book was released on 2016-11-03 with total page 448 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book encompasses the full breadth of the super-resolution imaging field, representing modern techniques that exceed the traditional diffraction limit, thereby opening up new applications in biomedicine. It shows readers how to use the new tools to increase resolution in sub-nanometer-scale images of living cells and tissue, which leads to new information about molecules, pathways and dynamics. The book highlights the advantages and disadvantages of the techniques, and gives state-of-the-art examples of applications using microscopes currently available on the market. It covers key techniques such as stimulated emission depletion (STED), structured illumination microscopy (SSIM), photoactivated localization microscopy (PALM), and stochastic optical reconstruction microscopy (STORM). It will be a useful reference for biomedical researchers who want to work with super-resolution imaging, learn the proper technique for their application, and simultaneously obtain a solid footing in other techniques.
Download or read book Single Molecule Spectroscopy written by R. Rigler and published by Springer Science & Business Media. This book was released on 2012-12-06 with total page 375 pages. Available in PDF, EPUB and Kindle. Book excerpt: The topics range from single molecule experiments in quantum optics and solid-state physics to analogous investigations in physical chemistry and biophysics.
Download or read book Lipid Uptake in Living Cells Visualized by Single Molecule Microscopy written by Sebastian Rhode and published by VDM Publishing. This book was released on 2008 with total page 116 pages. Available in PDF, EPUB and Kindle. Book excerpt: In recent years single molecule microscopy (SMM) became a very important tool in cell biology. Compared to other microscopic methods this technique offers the possibility to probe molecules in living cells with high temporal and lateral resolution. These advantages allow measuring the entire statistical distribution of a parameter. Therefore this method can be used to analyze heterogeneous probes, because it avoids the problems of averaging typically limiting investigations of ensembles. Another major advantage of this method, which is especially relevant for this work, is the possibility to determine the position of a labeled particle or molecule with accuracy which is about one order of magnitude below the diffraction limit of conventional optical microscopy. In the presented work all these benefits are utilized to answer questions concerning lipoprotein related lipid uptake and the behavior of lipids within the plasma membrane. Part of this work deals with the combination of atomic force microscopy (AFM) and fluorescence microscopy. The advantages of both methods are combined in one setup.
Download or read book Optical Fluorescence Microscopy written by Alberto Diaspro and published by Springer Science & Business Media. This book was released on 2010-11-25 with total page 252 pages. Available in PDF, EPUB and Kindle. Book excerpt: In the last decade, fluorescence microscopy has evolved from a classical “retrospective” microscopy approach into an advanced imaging technique that allows the observation of cellular activities in living cells with increased resolution and dimensions. A bright new future has arrived as the nano era has placed a whole new array of tools in the hands of biophysicists who are keen to go deeper into the intricacies of how biological systems work. Following an introduction to the complex world of optical microscopy, this book covers topics such as the concept of white confocal, nonlinear optical microscopy, fluctuation spectroscopies, site-specific labeling of proteins in living cells, imaging molecular physiology using nanosensors, measuring molecular dynamics, muscle braking and stem cell differentiation.
Download or read book Single Molecule Microscopy in Neurobiology written by Nobuhiko Yamamoto and published by Humana. This book was released on 2021-05-13 with total page 331 pages. Available in PDF, EPUB and Kindle. Book excerpt: This volume looks at the methodology and techniques used by experts to study how certain molecules function in specific locations, and their temporal patterns. Chapters in this book cover topics such as in vivo single-molecule tracking of voltage-gated calcium channels with split-fluorescent proteins in CRISPR-engineering C. elegans; protein-protein interactions in membranes using single particle tracking; neuropathological diseases revealed by quantum-dot single particle tracking; SPoD-OnSPAN; and investigating molecular diffusion inside small neuronal compartments with two-photon fluorescence correlation spectroscopy. In the Neuromethods series style, chapters include the kind of detail and key advice from the specialists needed to get successful results in your laboratory. Cutting-edge and comprehensive, Single Molecule Microscopy is a valuable resource for any researcher interested in learning more about this important field.
Download or read book Imaging and Spectroscopic Analysis of Living Cells written by and published by Academic Press. This book was released on 2012-01-25 with total page 573 pages. Available in PDF, EPUB and Kindle. Book excerpt: This volume of Methods in Enzymology is the first of three parts looking at current methodology for the imaging and spectroscopic analysis of live cells. The chapters provide hints and tricks not available in primary research publications. It is an invaluable resource for academics, researchers and students alike. - Expert authors who are leaders in the field - Extensively referenced and useful figures and tables - Provides hints and tricks to facilitate reproduction of methods
Download or read book Live Cell Imaging written by Robert D. Goldman and published by CSHL Press. This book was released on 2005 with total page 658 pages. Available in PDF, EPUB and Kindle. Book excerpt: Recent advances in imaging technology reveal, in real time and great detail, critical changes in living cells and organisms. This manual is a compendium of emerging techniques, organized into two parts: specific methods such as fluorescent labeling, and delivery and detection of labeled molecules in cells; and experimental approaches ranging from the detection of single molecules to the study of dynamic processes in organelles, organs, and whole animals. Although presented primarily as a laboratory manual, the book includes introductory and background material and could be used as a textbook in advanced courses. It also includes a DVD containing movies of living cells in action, created by investigators using the imaging techniques discussed in the book. The editors, David Spector and Robert Goldman, whose previous book was Cells: A Laboratory Manual,are highly respected investigators who have taught microscopy courses at Cold Spring Harbor Laboratory, the Marine Biology Laboratory at Woods Hole, and Northwestern University.
