| Contributors | p. ix |
| Preface | p. xiii |
| Acknowledgments | p. xv |
| Imaging Instrumentation | |
| High-Resolution Imaging of Cellular Processes in Caenorhabditis elegans | |
| Microscope Stand | p. 4 |
| Objective Lenses and Resolution | p. 5 |
| Transmitted Light Imaging | p. 8 |
| Fluorescence Imaging | p. 11 |
| Detectors | p. 13 |
| Confocal Imaging | p. 16 |
| Deconvolution Imaging | p. 19 |
| Whole Worm (Adult and Larval Stages) | p. 21 |
| Embryos | p. 25 |
| References | p. 33 |
| Immunofluorescence Microscopy | |
| Introduction | p. 36 |
| Preparing Specimens for Immunolocalization Studies | p. 42 |
| Preparation of Antibodies to Nematode Antigens | p. 54 |
| Microscopy: Necessary Equipment and Imaging Tips | p. 58 |
| Concluding Remarks | p. 60 |
| Recipes | p. 60 |
| References | p. 62 |
| Fluorescent Protein Methods: Strategies and Applications | |
| Introduction | p. 68 |
| Methods | p. 70 |
| Adopting New Fluorescent Proteins for Use in C. elegans | p. 80 |
| References | p. 89 |
| Modern Electron Microscopy Methods for C. elegans | |
| Introduction | p. 94 |
| Modern Electron Microscopes | p. 94 |
| Alternate Immersion Fixation Techniques | p. 97 |
| Serial Thin Sectioning | p. 116 |
| ImmunoEM Protocols | p. 124 |
| Alternate Methods to Post Stain Thin Sections | p. 126 |
| Electron Tomography | p. 130 |
| Emerging EM Technologies | p. 134 |
| Computer-Based Analytical Tools | p. 140 |
| Landmarks for Recognizing Tissues | p. 145 |
| Acknowledgments | p. 146 |
| References | p. 147 |
| Cell Biology | |
| Culture and Manipulation of Embryonic Cells | |
| Introduction: Uses and Limitations of the System | p. 154 |
| Embryo Devitellinization and Blastomere Isolation | p. 159 |
| Culture Methods | p. 163 |
| Drug Treatments | p. 165 |
| Radioactive Labeling | p. 166 |
| Fixation, Cytochemistry, and Electron Microscopy | p. 166 |
| Solutions and Culture Media | p. 170 |
| References | p. 174 |
| Laser Microsurgery in Caenorhabditis elegans | |
| Overview | p. 178 |
| Identifying Cells in C. elegans | p. 180 |
| Laser Ablation Theory and Apparatus | p. 186 |
| Laser Killing of Cells | p. 192 |
| Assessing Damage to the Operated Cell | p. 198 |
| Unintended Damage | p. 199 |
| Laser Cutting of Nerve Fibers | p. 199 |
| Related Methods | p. 201 |
| Acknowledgments | p. 203 |
| References | p. 203 |
| Methods in Cell Biology: Analysis of Cell Polarity in C. elegans Embryos | |
| Introduction | p. 208 |
| Methods for Analyzing Gene Function | p. 212 |
| Analyzing Cell Polarity: Protein and Organelle Localization | p. 215 |
| Dynamics of Polarity Establishment | p. 222 |
| Summary | p. 233 |
| References | p. 233 |
| Analysis of Membrane-Bound Organelles | |
| Introduction | p. 240 |
| Visualizing Organelles | p. 240 |
| Functional Studies | p. 250 |
| Protein Extract and Organelle Isolation | p. 259 |
| Summary | p. 261 |
| Acknowledgments | p. 261 |
| References | p. 262 |
| C. elegans Cell Cycle Analysis | |
| Introduction | p. 266 |
| Types of C. elegans cell cycles | p. 266 |
| Using the C. elegans Lineage to Follow Cell Divisions | p. 270 |
| Methods of Developmental Synchronization | p. 272 |
| Quantification of DNA Content | p. 275 |
| Detection of Different Cell Cycle Phases | p. 278 |
| Common cell Cycle Phenotypes and Potential Interpretations | p. 285 |
| References | p. 290 |
| -Physiology | |
| Methods for Studying Programmed Cell Death in C. elegans | |
| Introduction | p. 298 |
| Methods | p. 299 |
| Conclusions | p. 316 |
| Acknowledgments | p. 317 |
| References | p. 317 |
| Methods for Studying the DNA Damage Response in the Caenorhabdatis elegans Germ Line | |
| Introduction | p. 322 |
| Assay Development | p. 326 |
| Assays | p. 333 |
| Materials | p. 347 |
| Discussion | p. 348 |
| Acknowledgments | p. 349 |
| References | p. 349 |
| Analysis of Aging in Caenorhabditis elegans | |
| Introduction | p. 354 |
| Aging Pathways and Their Mechanism of Action | p. 355 |
| Life-Span Analysis | p. 358 |
| Analysis and Interpretation of Lifespan Data | p. 367 |
| Measurement of Age-Related Changes | p. 368 |
| Conclusion | p. 375 |
| Acknowledgments | p. 376 |
| References | p. 376 |
| Analyses of C. elegans Fat Metabolic Pathways | |
| Introduction | p. 384 |
| Staining of Fixed Animals and Histochemical Methods | p. 386 |
| Biochemical Methods | p. 390 |
| Biochemical Measurements of Metabolic Rates | p. 396 |
| Visualization of Fat Depots and Fat Metabolism Using Vital Dyes | p. 398 |
| Visualizing Hypodermal, Intestinal, and Gonad Lipid Stores using Nile Red | p. 400 |
| Fatty Acid Uptake | p. 403 |
| Raman Scattering-Based Methods of Lipid Imaging in C. elegans | p. 404 |
| Summary | p. 405 |
| Acknowledgements | p. 405 |
| References | p. 406 |
| Electrophysiological Methods for Caenorhabditis elegans Neurobiology | |
| Introduction | p. 410 |
| Recording Currents from Identified Neurons | p. 411 |
| Recording Synaptic Events from Identified Neurons | p. 417 |
| Recording Currents and Synaptic Events at the Neuromuscular Junction | p. 420 |
| Materials | p. 428 |
| Discussion | p. 433 |
| Summary | p. 433 |
| Acknowledgments | p. 434 |
| References | p. 434 |
| Index | p. 437 |
| Volumes in Series | p. 449 |
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