Description
A VALUABLE RESOURCE FOR INTRODUCING FLOW CYTOMETRY, PROVIDING A FOUNDATION FOR UNDERSTANDING ITS IMPORTANCE IN VARIOUS FIELDS
The newly updated and revised Flow Cytometry: First Principles, Third Edition covers flow cytometric theory and the fundamentals of instrumentation and data analysis using examples from a wide range of scientific and clinical applications. Readers will find and learn about both standard and advanced flow cytometric instrumentation, methods for analyzing flow data, and applications of the technique ranging from immunology, clinical diagnoses, DNA analysis, and cell sorting to cell function, plant biology, and molecular biology. Each chapter starts with basic knowledge and gradually expands on the subject matter, enabling readers to assess the ways that flow cytometry has been used in their respective fields and the ways that it might be applied in the future. A selected number of annotated references are given, aiming to point readers into relevant literature. There is an extensive glossary at the end of the book.
Written by two experts and highly illustrated, Flow Cytometry: First Principles, Third Edition includes information on:
- How flow cytometry is used to analyze cells
- How a flow cytometer can be used to separate different types of cells from each other
- Fluidics, electronics, and optics (including lasers, fluorochromes, and filters)
- Clinical applications of flow cytometry
An indispensable authoritative text on flow cytometry, the third edition of Flow Cytometry: First Principles is an essential resource for scientists, clinicians, technologists, instructors, and students.
Table of Contents
Preface xi
Acknowledgments: First Edition (A.L.G.) xv
Acknowledgments: Second Edition (A.L.G.) xvii
Acknowledgments: Third Edition (A.L.G. and P.K.W.) xix
1 The Past as Prologue 1
Further Reading 11
2 Setting the Scene 13
Shared Resource Laboratories 15
Data 16
Personnel 16
Training 17
Finances 19
Additional Thoughts 21
3 Instrumentation: Into the Black Box 23
Illumination of the Stream 24
Centering Cells in the Illuminating Beam: Hydrodynamic and Acoustic Focusing 26
Detection of Signals from Cells 33
Electronics 40
Further Reading 44
4 Information: Harnessing the Data 45
Data Storage 45
Software 48
Displaying Data 49
Analyzing Data 51
Further Reading 60
5 Seeing the Light: Lasers, Fluorochromes, Mirrors, Prisms, and Filters 61
General Theory 61
Lasers 63
Fluorochromes 66
Partitioning the Signal With Lenses, Filters, Mirrors, Prisms, and Gratings 77
Correcting for Spectral Overlap Between Fluorochromes 82
Further Reading 85
6 Cells from Without: Leukocytes, Surface Proteins, and the Strategy of Gating 87
Cells from Blood 88
Staining for Surface Proteins 93
Controls 96
Quantification of Fluorescence Intensity 101
Quantification of Volume and of Cell Number 104
Resolution 106
The Strategy of Gating 107
Gating On Fluorescence 114
Further Reading 117
7 Cells from Within: Intracellular Proteins 119
Methods for Permeabilizing Cells 121
Examples of Intracellular Staining 123
8 Cells from Within: DNA in Life and Death 127
Fluorochromes for DNA Analysis 127
Ploidy 130
Cell Cycle Analysis 136
Multicolor Analysis for DNA and Another Parameter 144
Chromosomes 151
Apoptosis and Necrosis 152
Autophagy 159
Further Reading 161
9 Assays of Function 163
Calcium Flux 164
Activation 166
Proliferation and Precursor Frequencies 168
Secretion of Soluble Analytes 171
Phagocytosis 173
Other Assays of Function 177
Comments 178
Further Reading 179
10 The Sorting of Cells 181
Flow Sorting: Theory and Practice 182
The Purity of Sorted Cells 188
Efficiency and Time 190
The Condition of Cells After Sorting 191
Small Sorters, "Simple" Sorters, Do-It-Yourself Sorting 193
Other Methods for Flow Sorting 195
Methods for Batch Sorting 196
Further Reading 198
11 Disease and Diagnosis: The Clinical Laboratory 199
Major Applications 201
Malaria and Flow Cytometry in the Wild 219
Sexing by Flow: Sorting Sperm for Good Reasons, for Bad Reasons, and for Profit 226
Comments 228
Further Reading 230
12 Out of the Mainstream: Biological Frontiers 231
Plants and Algae 232
Extracellular Vesicles 239
Fluorescent Proteins 240
Molecular Biology 242
Comments 245
Further Reading 245
13 Out of the Mainstream: Technological Frontiers 247
Mass Cytometers 247
Microfluidic Cytometers 249
Imaging Cytometers 251
In Summary 253
Further Reading 254
14 Flowing On: The Future 255
Artificial Intelligence and Robotics? 256
Individual or Shared? 257
Sophisticated Cytometers or Simple Cytometers? 258
How Many Fluorescence Parameters? 259
The Spirit of Flow Cytometry 259
Further Reading 260
Selected General Resources 263
Glossary 269
Figure Credits 297
Index 301
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