Light Microscopy

This volume addresses up-to-date light microscopy approaches and toolsets offered for live- or fixed-cell observations. The imaging strategies discussed in this book include confocal laser scanning and spinning disk confocal microscopy, FRET, FRAP, and laser microsurgery experiments. Chapters also describe the use of these imaging methodologies to study properties of a multitude of biomolecular targets in a broad range of model systems ranging from bacteria over tissue to whole animal imaging. Light Microscopy: Methods and Protocols puts special focus on system instrumentation parameters and sophisticated labeling and detection methods. 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, Light Microscopy: Methods and Protocols offers the novice user with straightforward strategies to address biological questions, while providing the experienced researcher with the latest applications that can be useful in routine practices. This book also serves as a useful teaching manual in practical courses on light microscopy.

Introduction to Modern Methods in Light Microscopy.- Three-Dimensional Live Imaging of Filamentous Fungi with Light Sheet-Based Fluorescence Microscopy (LSFM).- Light-Sheet Fluorescence Microscopy: Chemical Clearing and Labeling Protocols for Ultramicroscopy.- Two-Photon Intravital Microscopy Animal Preparation Protocol to Study Cellular Dynamics in Pathogenesis.- Imaging of Brain Slices with a Genetically Encoded Voltage Indicator.- FRET Microscopy for Real-Time Visualization of Second Messengers in Living Cells.- Imaging the Dynamics of Cell Wall Polymer Deposition in the Unicellular Model Plant, Penium margaritaceum.- Targeted Ablation Using Laser Nanosurgery.- Sample Preparation and Choice of Fluorophores for Single and Dual Color Photo-Activated Localization Microscopy (PALM) with Bacterial Cells.- STED Imaging in Drosophila Brain Slices.- Two-Color Total Internal Refection Fluorescence Microscopy of Exocytosis in Endocrine Cells.- Optical Coherence Microscopy.- Designing Image Analysis Pipelines in Light Microscopy: A Rationale Approach.- Automated Analysis of Intracellular Dynamic Processes.- Quantitative Image Analysis of Single-Molecule mRNA Dynamics in Living Cells.- Analysis of Protein Kinetics using Fluorescence Recovery After Photobleaching (FRAP).- Fluorescence Based High-Throughput and Targeted Image Acquisition and Analysis for Phenotypic Screening.