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The aim of this volume is to provide a comprehensive overview of optical tweezers setups, both in practical and theoretical terms, to help biophysicists, biochemists, and cell biologists to build and calibrate their own instruments and to perform force measurements on mechanoenzymes both in isolation in vitro and in living cells. Chapters have been divided in three parts focusing on theory and practical design of optical tweezers, detailed protocols for performing force measurements on single DNA- and microtubule/actin-associated mechanoenzymes in isolation, and describing recent advances that have opened up quantitative force measurements in living cells. 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.
Authoritative and cutting-edge, Optical Tweezers: Methods and Protocols aims help to further expand the accessibility and use of optical traps by scientists of diverse disciplines.
List of contents
Introduction to Optical Tweezers.- Exact Theory of Optical Tweezers and its Application to Absolute Calibration.- Beyond the Hookean Spring Model: Direct Measurement of Optical Forces Through Light Momentum Changes.- A Surfaced-Coupled Optical Trap with 1-bp Precision via Active Stabilization.- Implementation and Tuning of an Optical Tweezers Force-Clamp Feedback System.- Custom-Made Microspheres for Optical Tweezers.- Optical Torque Wrench Design and Calibration.- High-Resolution 'Fleezers': Dual-Trap Optical Tweezers Combined with Single-Molecule Fluorescence Detection.- Versatile Quadruple-Trap Optical Tweezers for Dual DNA Experiments.- Probing DNA-DNA Interactions with a Combination of Quadruple-Trap Optical Tweezers and Microfluidics.- Probing Single Helicase Dynamics on Long Nucleic Acids Through Force-Fluorescence Measurement.- Mechanically Watching the ClpXP Proteasome Machinery.- Deciphering the Molecular Mechanism of the Bacteriophage 29 DNA Packaging Motor.- Single-Molecule Protein Folding Experiments using High-Precision Optical Tweezers.- Observing Single RNA Polymerase Molecules Down to Base-Pair Resolution.- Optical Tweezers-Based Measurements of Forces Generated by Dynamic Microtubule Ends.- Simultaneous Manipulation and Super-Resolution Fluorescence Imaging of Individual Kinetochores Coupled to Microtubule Tips.- Measurement of Force-Dependent Release Rates of Cytoskeletal Motors.- Measuring The Kinetic and Mechanical Properties of Non-Processive Myosins using Optical Tweezers.- Quantifying Force and Viscoelasticity Inside Living Cells Using an Active-Passive Calibrated Optical Trap.- Measuring Molecular Forces using Calibrated Optical Traps in Living Cells.
Summary
The aim of this volume is to provide a comprehensive overview of optical tweezers setups, both in practical and theoretical terms, to help biophysicists, biochemists, and cell biologists to build and calibrate their own instruments and to perform force measurements on mechanoenzymes both in isolation in vitro and in living cells. Chapters have been divided in three parts focusing on theory and practical design of optical tweezers, detailed protocols for performing force measurements on single DNA- and microtubule/actin-associated mechanoenzymes in isolation, and describing recent advances that have opened up quantitative force measurements in living cells. 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.
Authoritative and cutting-edge, Optical Tweezers: Methods and Protocols aims help to further expand the accessibility and use of optical traps by scientists of diverse disciplines.
