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Cellular Electron Microscopy
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Cellular Electron Microscopy

McIntosh, J. Richard
ISBN-10: 0123706475
ISBN-13: 9780123706478

In our Marketplace:
2 new from $224.10
Recent advances in the imaging technique electron microscopy (EM) have improved the method, making it more reliable and rewarding, particularly in its description of three-dimensional detail. This book will help biologists from many disciplines understand modern EM and the value it might bring to their own work.
The books five sections deal with all major issues in EM of cells: specimen preparation, imaging in 3-D, imaging and understanding frozen-hydrated samples, labeling macromolecules, and analyzing EM data. Each chapter was written by scientists who are among the best in their field, and some chapters provide multiple points of view on the issues they discuss. Each section of the book is preceded by an introduction, which should help newcomers understand the subject. The book shows why many biologists believe that modern EM will forge the link between light microscopy of live cells and atomic resolution studies of isolated macromolecules, helping us toward the goal of an atomic resolution understanding of living systems. * Updates the numerous technological innovations that have improved the capabilities of electron microscopy * Provides timely coverage of the subject given the significant rise in the number of biologists using light microscopy to answer their questions and the natural limitations of this kind of imaging * Chapters include a balance of how to, so what and where next, providing the reader with both practical information, which is necessary to use these methods, and a sense of where the field is goingRecent improvements in electron microscopy of cells are bringing cell biologists back to "EM", particularly as they realize the limitations of the light microscopic methods that have been dominating the study of cell structure. This trend is a direct result of the resolving power of the electron microscope, of improved methods for specimen preparation, and of new technologies for three-dimensional imaging and image analysis. EM is beginning to fulfill its promise of linking light microscopy of live cells with atomic resolution studies of isolated macromolecules, with the goal of improving our understanding of living systems at atomic resolution. These new methods will help biologists from many disciplines to apply modern EM to their own work, allowing them to participate in the groundbreaking discoveries that are now being made in this field.A compilation of methods and protocols for doing electron microcopy of cells.
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Contributors
Introduction
Preparing Cells for Electron Microscopy
The Physics of Rapid Cooling and Its Implications for Cryoimmobilization of Cells
Introduction
Freezing Water
Biological Material
Freezing Biological Material
References
Cryopreparation Methods for Electron Microscopy of Selected Model Systems
Introduction
Equipment and Materials
General Rules for Loading Samples for HPF
Methods for Specific Organisms
Postfreezing Processing
Summary
References
Cryopreparation Methodology for Plant Cell Biology
Introduction
Rationale
Methods
Materials
Discussion
Concluding Remarks
References
Correlative Light and Electron Microscopy of Early Caenorhabditis elegans Embryos in Mitosis
Introduction
Rationale
Methods
Instrumentation and Materials
Discussion
References
Imaging Fixed Cells in Three Dimensions
Understanding Microtubule Organizing Centers by Comparing Mutant and Wild-Type Structures with Electron Tomography
Introduction
Rationale
Methods
Instrumentation and Materials
Discussion
Summary
References
Whole-Cell Investigation of Microtubule Cytoskeleton Architecture by Electron Tomography
Introduction
Rationale
Methods
Materials
Discussion
Summary
References
Electron Microscopy of Archaea
Introduction
Rationale
Methods and Materials
Discussion
Summary
References
Reconstructing Mammalian Membrane Architecture by Large Area Cellular Tomography
Introduction and Rationale
Methods and Materials
Discussion
Summary
References
Visualization of Membrane-Cytoskeletal Interactions During Plant Cytokinesis
Introduction
Rationale
Materials and Methods
Discussion
Summary
References
Electron Tomographic Methods for Studying the Chemical Synapse
Introduction and Rationale
Neurons, Neural Networks, and the Synapse
ET and the Synapse
Types of Neurons and Synapses
Sources of Neurons for Structural Studies
Methods for Assessing Neuron and Synapse Integrity
Rapid Freezing and HPF of Neurons
Modeling and Analysis of Synaptic Structures
Stimulation-Dependent Changes in Synaptic Structure
The Future of EM Tomography and Synapses
Conclusions
References
Using Electron Microscopy to Understand Functional Mechanisms of Chromosome Alignment on the Mitotic Spindle
Introduction
Basic Methodology
Organization of MTs in the Mitotic Spindle
Deciphering Mechanisms of Chromosome Alignment and K-Fiber Formation Using Correlative Microscopy
Toward a Molecular Understanding of Kinetochore Architecture
Beyond Morphology: Use of Electron Tomography to Model Kinetochore Control of MT Dynamics
Conclusions and Future Directions
References
Electron Microscopic Analysis of the Leading Edge in Migrating Cells
Introduction
Understanding the Mechanisms of Leading Edge Protrusion: Complementary Roles of LM and EM
Platinum Replica EM of the Leading Edge Cytoskeleton
Identification of Cytoskeletal Components in Platinum Replica Preparations
References
Imaging Actomyosin In Situ
Introduction and Overview
Preparing the Specimen
Data Collection and Tomogram Calculation
Identifying Similar Structures in an Ensemble
Building Atomic Models
Relating Class Averages to the Specimen
Application to IFM
Application to Other Types of Specimen
Summary
References
Imaging Frozen-Hydrated Cells and Cell Parts
Electron Tomography of Bacterial Chemotaxis Receptor Assemblies
Introduction
Analysis of Tsr Assemblies in Membrane Extracts
Electron Tomography of Fixed, Cryosectioned E. coli Cells
Cryo-Electron Microscopy of Frozen-Hydrated Sections of E. coli Cells
Structure of Chemotaxis Receptors Using Cryo-Electron Tomography
Summary
References
How to "Read" a Vitreous Section
Introduction
Conventional Versus Cryo-Electron Microscopy: A Comparison
Illustration: Two Pitfalls
References
Single-Particle Electron Cryomicroscopy of the Ion Channels in the Excitation-Contraction Coupling Junction
Introduction
Single-Particle Cryo-EM Methodology
3D Reconstruction of Proteins in Triad Junction
Spatial Arrangement of DHPRs and RyRs in Triad Junctions
Summary
References
Electron Microscopy of Microtubule-Based Cytoskeletal Machinery
Introduction
Rationale
Methods
Discussion
References
Reconstructing the Endocytotic Machinery
Introduction
Rationale
Methods
Results
Discussion
Summary
References
Localizing Macromolecules in Cells
3D Immunolocalization with Plastic Sections
Introduction
Rationale for the Various Approaches to Immuno-EM
Methods for Immuno-EM
Comparison of the Methods
Summary
References
Electron Microscopy Analysis of Viral Morphogenesis
Introduction
Analysis of Virus Assembly on Plastic Sections
Immunolabeling of Ultrathin Cryosections: Applications of the Tokuyasu Technique to Study Virus Assembly
Future Developments
Summary and Conclusions
References
Electron Tomography of Immunolabeled Cryosections
Introduction
Methods
Results and Discussion
References
Visualizing Macromolecules with Fluoronanogold: From Photon Microscopy to Electron Tomography
Introduction
Experimental Approach for Immunostaining of pKi-67 and RNAP I
Applications
Discussion
References
Markers for Correlated Light and Electron Microscopy
Introduction
How Do LM and EM Complement Each Other?
Fluorescence Photooxidation
Enzymatic-Based Methods
Particle-Based Methods for Protein Localization
Concluding Remarks
References
Localizing Specific Elements Bound to Macromolecules by EFTEM
Introduction
General Principles
Applications
Summary and Future Directions
References
Localization of Protein Complexes by Pattern Recognition
Introduction
Template Matching
The Missing-Wedge Problem
Applications
Conclusions
References
Aspects of Data Collection and Analysis
The Application of Energy-Filtered Electron Microscopy to Tomography of Thick, Selectively Stained Biological Samples
Introduction
Introduction to MPL Imaging
Detailed Description of MPL Tomography
Techniques
Results
Conclusions
References
Optimization of Image Collection for Cellular Electron Microscopy
Introduction: Assessing the Role of Image Collection in Cellular Electron Microscopy
Applications: The Importance and Meaning of SSNR in Cell Biology Applications
Optimization of Detectors and of the Detection Process for Maximization of SSNR
Detectors: Proven Technologies
Detectors: Experimental Technologies
Discussion
Conclusions
References
Future Directions for Camera Systems in Electron Microscopy
Introduction
Background and Rationale
Description of the Direct Detection Detector
Detector Characterization
Radiation Damage
Discussion
References
Structure Determination In Situ by Averaging of Tomograms
Introduction
Tomography and Its Increase in Resolution by Averaging
Coherent Averaging of Macromolecules in Practice
Applications of Tomogram Averaging
Conclusion and Outlook
References
Methods for Image Segmentation in Cellular Tomography
Introduction
Problem Formulation
Challenges in Cellular Tomography Segmentation
Important Considerations for Segmentation Methods
How Do We Know if the Segmentation Method Is Reliable?
Segmentation Methods for Cellular Tomography
Orientation-Based Segmentation
Results Using Orientation-Based Segmentation
Summary
References
Database Resources for Cellular Electron Microscopy
Introduction
The Cell Centered Database Project
Looking Ahead
References
Index
Volumes in Series

List price: $166.00
Edition: 2007
Publisher: Elsevier Science & Technology Books
Binding: Trade Cloth
Pages: 880
Size: 7.50" wide x 9.00" long x 1.25" tall
Weight: 3.78 lbs.
Language: English

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