Main description:

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.

Contents:

Preface...
Table of Contents...
Contributing Authors...

1. Introduction to Modern Methods in Light Microscopy

Part I Advanced Fluorescence Microscopy: Sample Preparation, Fluorophores and Model Systems

2. Three-Dimensional Live Imaging of Filamentous Fungi with Light Sheet-Based Fluorescence Microscopy (LSFM)
Francesco Pampaloni, Laura Knuppertz, Andrea Hamann, Heinz D. Osiewacz, and Ernst H.K. Stelzer

3. Light-Sheet Fluorescence Microscopy: Chemical Clearing and Labeling Protocols for Ultramicroscopy
Nina Jahrling, Klaus Becker, Saiedeh Saghafi, and Hans-Ulrich Dodt

4. Two-Photon Intravital Microscopy Animal Preparation Protocol to Study Cellular Dynamics in Pathogenesis
Erinke van Grinsven, Chloe Prunier, Nienke Vrisekoop, and Laila Ritsma

5. Imaging of Brain Slices with a Genetically Encoded Voltage IndicatorPeter Quicke, Samuel J. Barnes, and Thomas Knoepfel

6. FRET Microscopy for Real-Time Visualization of Second Messengers in Living Cells
Axel E. Kraft and Viacheslav O. Nikolaev

7. Imaging the Dynamics of Cell Wall Polymer Deposition in the Unicellular Model Plant, Penium margaritaceum
David Domozych, Anna Lietz, Molly Patten, Emily Singer, Berke Tinaz, and Sandra C. Raimundo

8. Targeted Ablation Using Laser Nanosurgery
Naga Venkata Gayathri Vegesna, Paolo Ronchi, Sevi Durdu, Stefan Terjung, and Rainer Pepperkok

Part II Super and High-Resolution Optical Imaging

9. Sample Preparation and Choice of Fluorophores for Single and Dual Color Photo-Activated Localization Microscopy (PALM) with Bacterial Cells
Juri N. Bach, Giacomo Giacomelli, and Marc Bramkamp

10. STED Imaging in Drosophila Brain Slices
Sandra Fendl, Jesus Pujol-Marti, Joel Ryan, Alexander Borst, and Robert Kasper

11. Two-Color Total Internal Refection Fluorescence Microscopy of Exocytosis in Endocrine Cells
Adam J. Trexler and Justin W. Taraska

12. Optical Coherence Microscopy
Rainer A. Leitgeb

Part III Quantitative and Computational Image Analysis

13. Designing Image Analysis Pipelines in Light Microscopy: A Rationale Approach
Ignacio Arganda-Carreras and Philippe Andrey

14. Automated Analysis of Intracellular Dynamic Processes
Yao Yao, Ihor Smal, Ilya Grigoriev, Maud Martin, Anna Akhmanova, and Erik Meijering

15. Quantitative Image Analysis of Single-Molecule mRNA Dynamics in Living Cells
Jose Rino, Ana C. de Jesus, and Maria Carmo-Fonseca

16. Analysis of Protein Kinetics using Fluorescence Recovery After Photobleaching (FRAP)
Nickolaos-Nikiforos Giakoumakis, Maria Anna Rapsomaniki, and Zoi Lygerou

17. Fluorescence Based High-Throughput and Targeted Image Acquisition and Analysis for Phenotypic Screening
Manuel Gunkel, Jan Philipp Eberle, and Holger Erfle