Cellular imaging techniques for neuroscience and beyond / [edited by] Floris G. Wouterlood. — London ; Waltham, MA : Elsevier/Academic Press, 2012. – (64.118/C393) |
Contents
CONTENTS
List of Contributors
Chapter 1 Confocal Laser Scanning: of Instrument, Computer Processing, and Men 1
Introduction 2
Pinhole, Depth of Focus, and Laser Illumination 2
When/Why Does One Need a CLSM? 4
Abbe, Shannon, and Nyquist 8
Imaging of a 2D Line and Deblurring 9
Axial Resolution 11
Resolution and Sampling 12
Signal Separation, Orders of Magnitude, and Resolution Limits 13
Confocal Microscopy Further Considered 14
Cross Talk Awareness 16
Elimination of Cross Talk 18
Biological Objects Translated to Pixels 19
High-probability Determination of Diameter 20
Why Does a 3D Reconstructed Cell Resemble a Pancake? 24
Touch 25
Actual Experiment 26
Synaptic Contacts: Extra Marker 29
Colocalization 30
Conclusion 32
Acknowledgments 32
References 33
Chapter 2 Beyond Abbe's Resolution Barrier: STED Microscopy 35
Introduction 36
A New Wave of Imaging 37
STED Microscopy: The Basic Concept 39
Implementation of STED Microscopy
Sine Qua Non: Speed, Color, Depth, Live Imaging
Summary and Outlook
References
Chapter 3 Enhancement of Optical Resolution by 4pi Single and Multiphoton Confocal Fluorescence Microscopy 55
Introduction 56
The 4pi Principle and Setup 56
Microscope Alignment 58
4pi Imaging 60
4pi Deconvolution 61
Sample Preparation 62
Microtubule and Microtubule Plus End Imaging 65
Visualization of DNA 66
Single-photon Excitation (Measurement of the Redox State in Dopamine Neurons) 68
SYCP3 Axis as a Marker for Chromatin Organization in Mouse Spermatocytes 70
Microbubbles with Medicine 75
Future of 4pi Imaging 77
Acknowledgment 77
References 77
Chapter 4 Nano Resolution Optical Imaging Through Localization Microscopy 81
Introduction 82
Superresolution Microscopy Techniques 82
The Main Approaches to Single-molecule Localization-based Superresolution Microscopy 87
Fluorescent Probes 89
Fluorescent Proteins 89
Multicolor Localization Microscopy 90
Outlook 93
Conclusion 96
Acknowledgments 96
References 97
Chapter 5 Optical Investigation of Brain Networks Using Structured Illumination 101
Introduction 101
Structuring Light by Phase Modulation Using SLMs 103
Wavefront Engineering Using SLMs: The Optical Setup 104
Light-sensitive Molecular Tools for the Investigation of the Central Nervous System 111
SLM-based Approaches for the Optical Dissection of Brain Microcircuits 112
Conclusions 116
Acknowledgments 116
References 116
Chapter 6 Multiphoton Microscopy Advances Toward Super Resolution 121
Introduction 121
Point Spread Function for Single- and Multiphoton Imaging 123
Super Resolution Techniques for Multiphoton Fluorescence Microscopy 126
Conclusions 134
Acknowledgments 134
References 134
Chapter 7 The Cell at Molecular Resolution: Principles and Applications of Cryo-Electron Tomography 141
Introduction: Cellular Landscapes at Molecular Resolution 141
The Cryo-ET Method 143
Detection, Identification, and Hybrid Methods 151
Conclusions 175
Acknowledgments 178
References 178
Chapter 8 Cellular-Level Optical Biopsy Using Full-Field Optical Coherence Microscopy 185
Introduction 185
The FF-OCM Technique 187
Detection Sensitivity 189
Spatial Resolution 191
Sample Motion Artifacts 192
FF-OCM for High-Resolution "Optical Biopsy" 193
Conclusion 195
Acknowledgment 196
References 196
Chapter 9 Retroviral Labeling and Imaging of Newborn Neurons in the Adult Brain. 201
Techniques to Label and Detect Newborn Neurons in the Adult Brain 202
Retrovirus-mediated Labeling of Adult-born Neurons 203
Single-cell Genetic Manipulation in Adult-born Neurons 205
Retrovirus Production and Delivery 207
Viral-labeled Cell Toxicity and Physiological Changes 209
Imaging Newborn Neurons in the Adult Brain 210
In Vivo Live Animal Imaging of Adult Neurogenesis 210
In Vivo Window Preparation 211
In Vivo Imaging Setup and Acquisition 213
Postacquisition Image Processing and Analysis 213
Future Directions in Live Animal Imaging of Adult Neurogenesis 214
Acknowledgments 216
References 216
Chapter 10 Study of Myelin Sheaths by CARS Microscopy 221
Traditional Myelin Imaging Methods 221
Principle and History of CARS Microscopy 224
Technical Characteristics of CARS Microscopy 226
CARS Microscopy for Ex Vivo and In Vivo Myelin Imaging 227
Mechanistic Understanding of Demyelination and Remyelination Enabled by CARS Imaging 231
Other Methods for In Vivo Imaging of Myelin 236
Outlook for Myelin Imaging by CARS Microscopy 238
Acknowledgments 240
References 240
Chapter 11 High-Resolution Approaches to Studying Presynaptic Vesicle Dynamics Using Variants of FRAP and Electron Microscopy 247
Introduction 248
Quantifying Dynamic Events at the Macromolecular Scale 249
FRAP for Studying Mobility 251
Variations on FRAP Using Photoswitchable Fluorophores 255
Linking Fluorescence and Ultrastructure: Correlative Approaches for Assaying Presynaptic Function 259
Structure-Function Relationships of Vesicle Pools in Hippocampal Synapses 263
Combining FRAP with Correlative Electron Microscope 265
Concluding Remarks 269
Acknowledgments 269
References 270
Index 275
Color Plates