Cytoskeleton Methods and Protocols

In the ten years since the publication of the first edition, great advances in fluorescent labeling, optics, and sample preparation have significantly improved the imaging capability of microscopy, allowing for a continual refinement of our understanding of the cytoskeleton as a dynamic synergy of components. In Cytoskeleton Methods and Protocols, Second Edition, internationally renowned experts present techniques which reflect many of the recent technological advances in experimental tools for cytoskeleton research with emphasis on animal, plant, protist, and fungal model systems. This cutting-edge volume contains methods for live-cell imaging, fluorescence microscopy, electron microscopy, analysis of cell and organelle motility, isolation of cytoskeleton components, and proteomics, amongst other topics. As a volume in the highly successful Methods in Molecular Biology™ series, chapters incorporate introductions to their respective subjects, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and notes that provide unpublished technical information on troubleshooting and avoiding known pitfalls.

Up-to-date and comprehensive, Cytoskeleton Methods and Protocols, Second Edition serves as an ideal guide to scientists who wish to continue this fruitful and important biological research.

Part I: Live-Cell Imaging of the Cytoskeleton 1. High Resolution Multimode Light Microscopy of Cell Migration: Long-Term Imaging and Analysis Torsten Wöllert and George M. Langford 2. Imaging the Cytoskeleton in Live Xenopus laevis Embryos Sarah Woolner, Ann L. Miller, and William M. Bement 3. Live-Cell Imaging of the Cytoskeleton and Mitochondrial-Cytoskeletal Interactions in Budding Yeast Theresa C. Swayne, Thomas G. Lipkin, and Liza A. Pon Part II: Dynamics of Actin Filaments and Microtubules 4. Jasplakinolide: An Actin-Specific Reagent that Promotes Actin Polymerization Andreas Holzinger 5. Analysis of Centrosome Function and Microtubule Dynamics by Time-Lapse Microscopy in Xenopus Egg Extracts Christiane Wiese, Jonathan R. Mayers, and Alison J. Albee Part III: Fluorescence Microscopy Protocols for Investigating Cytoskeleton Components 6. Quantitative Fluorescence Microscopy Techniques Alessandro Esposito, Simon Schlachter, Gabriele S. Kaminski Schierle, Alan D. Elder, Alberto Diaspro, Fred S. Wouters, Clemens F. Kaminski, and Asparouh I. Iliev 7. A FRET-Based Approach for Studying Conformational Changes of a Cytoskeleton-Related Tumor Suppressor Molecule Robert F. Hennigan, Mary F. Chaiken, Lauren A. Foster, and Wallace Ip 8. Sample Preparation for Fluorescence Imaging of the Cytoskeleton in Fixed and Living Plant Roots Julia Dyachok, Cheol-Min Yoo, Karuppaiah Palanichelvam, and Elison B. Blancaflor 9. Imaging of the Cytoskeleton and Mitochondria in Fixed Budding Yeast Cells Theresa C. Swayne, Istvan R. Boldogh, and Liza A. Pon Part IV: Electron Microscopy Protocols for Imaging the Cytoskeleton 10. Imaging Cytoskeleton Components by Electron Microscopy Tatyana Svitkina 11.Purification and Localization of Intraflagellar Transport Particles and Polypeptides Roger D. Sloboda 12. High Pressure Freezing, Electron Microscopy, and Immuno-Electron Microscopy of Tetrahymena thermophila Basal Bodies Janet B. Meehl, Thomas H. Giddings, Jr., and Mark Winey 13. Strategies for Imaging Microtubules in Plant Cells Andreas Holzinger, Eiko Kawamura, and Geoffrey O. Wasteneys Part V: Analysis of Cell and Organelle Motility 14. A Rapid Tracking Method for the Quantitative Analysis of Organelle Streaming Velocity Carola Luise Holweg 15. Melanosome Motility in Fish Retinal Pigment Epithelial (RPE) Cells Christina King-Smith 16. Analysis of Properties of Cilia Using Tetrahymena thermophila Vidyalakshmi Rajagopalan, Elizabeth O. Corpuz, Mark J. Hubenschmidt, Caroline R. Townsend, David J. Asai, and David E. Wilkes 17. Application of Cell Traction Force Microscopy for Cell Biology Research James H-C. Wang and Bin Li 18. 2D and 3D Quantitative Analysis of Cell Motility and Cytoskeletal Dynamics Deborah Wessels, Spencer Kuhl, and David R. Soll 19. Detergent-Extracted Models for the Study of Cilia or Flagella Charles B. Lindemann and Kathleen A. Lesich Part VI: Genetic and Proteomics Approaches for Cytoskeleton Protein Function 20. The Analysis of Intermediate Filament Dynamics Using Transfections and Cell Fusions Jesús M. Paramio 21. Functional Analysis of Actin-Binding Proteins in the Central Nervous System of Drosophila Qi He 22. Proteomic Tools for the Analysis of Cytoskeleton Proteins Ewa Wywial, Vishnubhat Nataraj Dongre, and Shaneen M. Singh