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Recent advances in single molecule science have presented a new branch of science: single molecule cellular biophysics, combining classical cell biology with cutting-edge single molecule biophysics. This textbook explains the essential elements of this new discipline, from the state-of-the-art single molecule techniques to real-world applications in unravelling the inner workings of the cell. Every effort has been made to ensure the text can be easily understood by students from both the physical and life sciences. Mathematical derivations are kept to a minimum whilst unnecessary biological terminology is avoided and text boxes provide readers from either background with additional information. 100 end-of-chapter exercises are divided into those aimed at physical sciences students, those aimed at life science students and those that can be tackled by students from both disciplines. The use of case studies and real research examples make this textbook indispensable for undergraduate students entering this exciting field.
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.
This third edition volume expands on the previous editions with new discussions on the latest techniques and developments in the field. The chapters in this book are organized into four parts, and cover topics such as optical tweezers; single-molecule fluorescence tools; atomic force microscopy; magnetic tweezers; applications to virus protein shells, unfolding of proteins, nucleic acids, motor proteins, in vivo and in vitro; and protocols to establish specific surface interactions and perform force calibration. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Cutting-edge and thorough, Single Molecule Analysis: Methods and Protocols, Third Edition is a valuable resource for all researchers who want to learn more about this exciting and still expanding field. Chapters 2, 7, 8, 9, 12, 18, and 19 are available open access under a Creative Commons Attribution 4.0 International License via link.springer.com.
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.
Single-Molecule Enzymology, Part B, the latest volume in the Methods in Enzymology series, continues the legacy of this premier serial with quality chapters authored by leaders in the field. This volume covers research methods in single-molecule enzymology, and includes sections on such topics as force-based and hybrid approaches, fluorescence, high-throughput sm enzymology, and nanopore and tethered particle motion. - Continues the legacy of this premier serial with quality chapters authored by leaders in the field - Covers research methods in single-molecule enzymology - Contains sections on such topics as force-based and hybrid approaches, fluorescence, high-throughput sm enzymology, and nanopore and tethered particle motion
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
This detailed volume explores a wide variety of techniques involving optical tweezers, a technology that has become increasingly more accessible to a broad range of researchers. Beginning with recent technical advances, the book continues by covering the application of optical tweezers to study DNA-protein interactions and DNA motors, protocols to perform protein (un)folding experiments, the application of optical tweezers to study actin- and microtubule-associated motor proteins, and well as protocols for investigating the function and mechanical properties of microtubules and intermediate filaments, and more. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Optical Tweezers: Methods and Protocols, Second Edition serves as an ideal resource for expanding the accessibility and use of optical traps by scientists of diverse disciplines.
Thanks to the pioneering works of Ashkin and coworkers, optical tweezers (OTs) have become an invaluable tool for myriad studies throughout the natural sciences. Their success relies on the fact that they can be considered as exceptionally sensitive transducers that are able to resolve pN forces and nm displacements, with high temporal resolution, down to μs. Hence their application to study a wide range of biological phenomena such as measuring the compliance of bacterial tails, the forces exerted by a single motor protein, and the mechanical properties of human red blood cells and of individual biological molecules. The number of articles related to them totals to a whopping 58,000 (source Google Scholar)! Microrheology is a branch of rheology, but it works at micrometer length scales and with microliter sample volumes. Therefore, microrheology techniques have been revealed to be very useful tools for all those rheological/mechanical studies where rare or precious materials are employed, such as in biological and biomedical studies. The aim of this book is to provide a pedagogical introduction to the physics principles governing both the optical tweezers and their application in the field of microrheology of complex materials. This is achieved by following a linear path that starts from a narrative introduction of the "nature of light," followed by a rigorous description of the fundamental equations governing the propagation of light through matter. Moreover, some of the many possible instrumental configurations are presented, especially those that better adapt to perform microrheology measurements. In order to better appreciate the microrheological methods with optical tweezers explored in this book, informative introductions to the basic concepts of linear rheology, statistical mechanics, and the most popular microrheology techniques are also given. Furthermore, an enlightening prologue to the general applications of optical tweezers different from rheological purposes is provided at the end of the book.
The technical development of optical tweezers, along with their application in the biological and physical sciences, has progressed significantly since the demonstration of an optical trap for micron-sized particles based on a single, tightly focused laser beam was first reported more than twenty years ago. Bringing together many landmark papers on