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Summary
Summary
Correlative Light and Electron Microscopy III, Volume 140 , a new volume in the Methods in Cell Biology series, continues the legacy of this premier serial with quality chapters authored by leaders in the field. Topics discussed in this new release include Millisecond time-resolved CLEM, Super resolution LM und SEM of high-pressure frozen C. elegans, Preservation fluorescence, super res CLEM, APEX in Tissue, Corrsight mit IBIDI flowthrough chamber, Correlative Light Atomic Force Electronic Microscopy (CLAFEM), Atmospheric EM CLEM, and High-precision correlation, amongst other topics.
Chapters in this ongoing series deal with different approaches for analyzing the same specimen using more than one imaging technique. The strengths and application area of each is presented, with this volume exploring the aspects of sample preparation of diverse biological systems for different CLEM approaches.
Author Notes
Dr. Thomas M#65533;ller-Reichert is interested in how the microtubule cytoskeleton is modulated within cells to fulfill functions in meiosis, mitosis and abscission. The M#65533;ller-Reichert lab is mainly applying correlative light microscopy and electron tomography to study the 3D organization of microtubules in the early embryo of the nematode Caenorhabditis elegans and in tissue culture cells. He got his PhD degree from the Swiss Federal Institute of Technology (ETH) in Zurich and moved afterwards to the EMBL in Heidelberg (Germany) for a post-doc with Dr. Tony Hyman. He was a visiting scientist with Dr. Kent McDonald (UC Berkeley, USA) and set up the electron microscope facility at the newly founded Max Planck Institute of Molecular Cell Biology and Genetics (MPI-CBG). Since 2010 he is head of the Core Facility Cellular Imaging (CFCI) of the Medical Faculty Carl Gustav Carus of the TU Dresden (Germany).
Together with Dr. Paul Verkade he has developed a rapid transfer system for high-pressure freezing used for Correlative Light and Electron Microcopy. He has organized a number microscopy conferences and taught in several (CL)EM courses. He edited an MCB volume on the Electron Microscopy of Model Systems.
Dr. Paul Verkade's research focuses on the sorting mechanisms in intracellular transport pathways. His main tools are microscopy techniques, with an emphasis on electron microscopy (EM) in which field he has published over 50 papers. He has studied and got his PhD degree at the University of Utrecht, the Netherlands. After his post-doc time at the EMBL, Heidelberg, Germany in the group of Kai Simons and setting up a new EM lab at the Max Planck Institute for Molecular Cell Biology in Dresden, Germany he moved to the University of Bristol, UK in 2006. Here he set up a new EM unit as part of the Wolfson Bioimaging Facility, a fully integrated light and electron microscopy centre.
To support his transport studies, part of his research is to develop techniques and tools for the use of Correlative Light Electron Microscopy (CLEM). Amongst other things he has developed the Rapid Transfer System for the EMPACT2 high-pressure freezer together with Leica Microsystems. This allows for the combination of time-resolved CLEM with optimal preservation of ultrastructure for EM.
Dr. Verkade is chair of the Electron Microscopy section of the Royal Microscopical Society (RMS) and of the Cryo Microscopy Group, affiliated to the RMS. He has organised and taught on a large number of courses and workshops on subjects such as high-pressure freezing, Correlative Light Electron Microscopy, and immuno EM. He is also the principle organiser of the EMBO practical course on CLEM.
Table of Contents
Contributors | p. xi |
Preface | p. xvii |
Chapter 1 Millisecond Time Resolution Correlative Light and Electron Microscopy for Dynamic Cellular Processes | p. 1 |
Introduction | p. 2 |
1 Methods | p. 3 |
2 Instrumentation and Materials | p. 15 |
3 Discussion and Outlook | p. 17 |
Acknowledgments | p. 18 |
References | p. 18 |
Chapter 2 3D Subcellular Localization With Superresolution Array Tomography on Ultrathin Sections of Various Species | p. 21 |
Introduction and Rationale | p. 22 |
1 Methods-Core Protocol | p. 23 |
2 Methods-Model-Specific Adaptations and Considerations | p. 34 |
3 Instrumentation and Materials | p. 41 |
Conclusions | p. 43 |
Acknowledgments | p. 45 |
References | p. 45 |
Chapter 3 Preserving the Photoswitching Ability of Standard Fluorescent Proteins for Correlative In-Resin Super-resolution and Electron Microscopy | p. 49 |
Introduction | p. 50 |
1 Rationale | p. 52 |
2 Materials | p. 54 |
3 Methods | p. 55 |
4 Discussion | p. 63 |
Acknowledgments | p. 65 |
References | p. 65 |
Chapter 4 Minimal Resin Embedding of Multicellular Specimens for Targeted FIB-SEM Imaging | p. 69 |
Introduction | p. 70 |
1 Rationale | p. 71 |
2 Methods | p. 71 |
3 Results: FIB-SEM Targeting and Imaging | p. 75 |
4 Instrumentation and Materials | p. 77 |
5 Discussion | p. 79 |
Acknowledgments | p. 81 |
References | p. 81 |
Chapter 5 A New Method for Cryo-Sectioning Cell Monolayers Using a Correlative Workflow | p. 85 |
Introduction | p. 86 |
1 Rationale | p. 87 |
2 Methods | p. 88 |
3 Instrumentation, Materials, and Reagents | p. 98 |
4 Results and Discussion | p. 100 |
Acknowledgments | p. 101 |
References | p. 101 |
Chapter 6 Correlative Light and Electron Microscopic Detection of GFP-Labeled Proteins Using Modular APEX | p. 105 |
1 Vectors for Correlative Light and Electron Microscopy in Mammalian Cell Culture and Whole Zebrafish | p. 107 |
2 Correlative Light and Electron Microscopy in Cell Culture | p. 108 |
3 Subcellular Protein Distribution Analysis of Transgenic Zebrafish | p. 115 |
References | p. 121 |
Chapter 7 Correlation of Live-Cell Imaging With Volume Scanning Electron Microscopy | p. 123 |
Introduction | p. 124 |
1 Rationale | p. 126 |
2 Methods | p. 126 |
3 Materials | p. 139 |
4 Discussion | p. 140 |
Conclusions | p. 145 |
Acknowledgments | p. 146 |
References | p. 146 |
Chapter 8 A Fully Integrated, Three-Dimensional Fluorescence to Electron Microscopy Correlative Workflow | p. 149 |
Introduction | p. 150 |
1 Materials and Methods | p. 151 |
2 Discussion | p. 159 |
3 Summary | p. 161 |
Acknowledgments | p. 161 |
Supplementary data | p. 162 |
References | p. 162 |
Chapter 9 CLAFEM: Correlative Light Atomic Force Electron Microscopy | p. 165 |
Introduction | p. 166 |
1 Materials and Methods | p. 170 |
2 Results and Discussion | p. 173 |
Acknowledgments | p. 181 |
References | p. 181 |
Chapter 10 Correlative Light-Electron Microscopy in Liquid Using an Inverted SEM (ASEM) | p. 187 |
Introduction | p. 188 |
1 Instrument Design and Sample Geometry of the Atmospheric Scanning Electron Microscopy | p. 189 |
2 The Application of Correlative Light-Electron Microscopy Using Atmospheric Scanning Electron Microscopy | p. 193 |
3 Discussion | p. 208 |
Conclusion | p. 210 |
Acknowledgments | p. 211 |
References | p. 211 |
Chapter 11 Relocation is the Key to Successful Correlative Fluorescence and Scanning Electron Microscopy | p. 215 |
Introduction | p. 216 |
1 Rationale | p. 219 |
2 Methods | p. 220 |
3 Instrumentation and Materials | p. 228 |
4 Results | p. 230 |
5 Discussion | p. 235 |
Acknowledgments | p. 240 |
References | p. 240 |
Chapter 12 Correlative Two-Photon and Serial Block Face Scanning Electron Microscopy in Neuronal Tissue Using 3D Near-Infrared Branding Maps | p. 245 |
Introduction | p. 246 |
1 Rationale | p. 251 |
2 Methods | p. 251 |
Concluding Remarks | p. 273 |
Acknowledgments | p. 274 |
References | p. 274 |
Chapter 13 Find Your Way With X-Ray: Using microCT to Correlate In Vivo Imaging With 3D Electron Microscopy | p. 277 |
Introduction | p. 278 |
1 Methods | p. 280 |
2 Instrumentation and Materials | p. 297 |
3 Discussion | p. 298 |
Acknowledgments | p. 299 |
References | p. 299 |
Chapter 14 triCLEM: Combining High-Precision, Room Temperature CLEM With Cryo-Fluorescence Microscopy to Identify Very Rare Events | p. 303 |
Introduction | p. 304 |
1 Methods | p. 307 |
2 Instrumentation and Materials | p. 315 |
3 Discussion | p. 316 |
Acknowledgments | p. 317 |
References | p. 318 |
Chapter 15 Matrix MAPS-An Intuitive Software to Acquire, Analyze, and Annotate Light Microscopy Data for CLEM | p. 321 |
Introduction | p. 322 |
1 The Software | p. 323 |
2 Workflow | p. 325 |
3 Discussion/Outlook | p. 330 |
Acknowledgments | p. 331 |
References | p. 331 |
Chapter 16 eC-CLEM: a Multidimension, Multimodel Software to Correlate Intermodal Images With a Focus on Light and Electron Microscopy | p. 335 |
Introduction | p. 336 |
1 Material | p. 336 |
2 Image Preprocessing and Automatic Detection of the Melanosomes in Both Imaging Modalities | p. 338 |
3 Identify the Transformation Project | p. 343 |
4 Setting the Autofinder Parameters to Get Accurate Initial Registration | p. 346 |
5 Manual Registration Versus Autofinder | p. 350 |
6 3D Transformations | p. 351 |
References | p. 351 |