Alice Longobardi Givan The Herbert C. Englert Cell Analysis Laboratory of the Norris Cotton Cancer Center and Department of Physiology Dartmouth Medical School Lebanon, New Hampshire Wiley-Liss A John Wiley & Sons, Inc., Publication 2001 FROM BACK COVER: Flow cytometry continually amazes scientists with its ever-expanding utility. Advances in flow cytometry have opened new directions in theoretical science, clinical diagnosis, and medical practice. The new edition of Flow Cytometry: First Principles provides a thorough update of this now classic text, reflecting innovations in the field while outlining the fundamental elements of instrumentation, sample preparation, and data analysis. Flow Cytometry: First Principles, Second Edition explains the basic principles of flow cytometry, surveying its primary scientific and clinical applications and highlighting state-of-the-art techniques at the frontiers of research. This edition contains extensive revisions of all chapters, including new discussions on fluorochrome and laser options for multicolor analysis, an additional section on apoptosis in the chapter on DNA, and new chapters on intracellular protein staining and cell sorting, including high-speed sorting and alternative sorting methods, as well as traditional technology. This essential resource: * Assumes no prior knowledge of flow cytometry * Progresses with an informal, engaging lecture style from simple to more complex concepts * Offers a clear introduction to new vocabulary, principles of instrumentation, and strategies for data analysis * Emphasizes the theory relevant to all flow cytometry, with examples from a variety of clinical and scientific fields Flow Cytometry: First Principles, Second Edition provides scientists, clinicians, technologists, and students with the knowledge necessary for beginning the practice of flow cytometry and for understanding related literature. CONTENTS Preface xi Acknowledgments (First Edition) xv Acknowledgments (Second Edition) xvii 1. The Past as Prologue 1 2. Setting the Scene 11 3. Instrumentation: Into the Black Box 15 Illumination of the Stream 16 Centering Cells in the Illuminating Beam 18 Detection of Signals from Cells 25 Electronics 31 4. Information: Harnessing the Data 41 Data Storage 41 Data Analysis 43 5. Seeing the Light: Lasers, Fluorochromes, and Filters 59 General Theory 59 Lasers 61 Fluorochromes 65 Partitioning the Signal with Lenses, Filters, and Mirrors 72 Spectral Compensation 76 6. Cells from Without: Leukocytes, Surface Proteins, and the Strategy of Gating 81 Cells from Blood 82 Staining for Surface Markers 87 Controls 90 Quantitation 95 Sensitivity 98 The Strategy of Gating 99 Gating on Fluorescence 108 7. Cells from Within: Intracellular Proteins 115 Methods for Permeabilizing Cells 116 Examples of Intracellular Staining 118 8. Cells from Within: DNA in Life and Death 123 Fluorochromes for DNA Analysis 123 Ploidy 125 Cell Cycle Analysis 131 Two-Color Analysis for DNA and Another Parameter 142 Chromosomes 147 Apoptosis 150 Necrosis 154 9. The Sorting of Cells 159 Sorting Theory 159 Characterization of Sorted Cells 166 Alternative Methods for Sorting 170 The Condition of Cells After Sorting 172 10. Disease and Diagnosis: The Clinical Laboratory 175 The Hematology Laboratory 178 The Pathology Laboratory 186 Solid Organ Transplantation 189 Comments 192 11. Out of the Mainstream: Research Frontiers 195 Functional Assays 196 The Aquatic Environment 202 Reporter Molecules 207 Microbiology 211 Molecular Biology 213 Multiplex Cytometry for Soluble Analytes 218 Reproductive Technology 220 12. Flowing On: The Future 225 General References 229 Glossary 235 Figure Credits 257 Index 263 WHERE TO ORDER: John Wiley & Sons, Inc. Distribution Center 1 Wiley Drive Somerset, NJ 08875-1272 Phone: (732) 469-4400 or (800) 225-5945 Fax: (732) 302-2300 E-mail: bookinfo@wiley.com PRICE: $44.95 (paperback) ISBN: 0-471-38224-8
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