Cytometry, Part A: Wilson; Matsudaira; Darzynkiewicz; Robinson; Crissman: 9780122030536: Book: Science: eCampus.com

Each chapter presents a detailed background of the described method, its theoretical foundations, and its applicability to different biomedical material. Updated chapters describe either the most popular methods or those processes that have evolved the most since the past edition. Additionally, a large portion of the volume is devoted to clinical cytometry. Particular attention is paid to applications of cytometry in oncology, the most rapidly growing area.

Key Features
* Contains 56 extensive chapters authored by world authorities on cytometry
* Covers a wide range of topics, including principles of cytometry and general methods, cell preparation, tandardization and quality assurance, cell proliferation, apoptosis, cell-cell/cell-environmental interactions, cytogenetics and molecular genetics, cell function and differentiation, experimental and clinical oncology, microorganisms, and infectious diseases
* Describes in-depth the essential methods and scientific principles of flow and laser scanning cytometry and illustrates how they can be applied to the fields of biology and medicine
* Complements the first and second editions on flow cytometry in the Methods in Cell Biology series and includes new sections on technology principles.
New York Medical College, Hawthorne. Presents a detailed discussion of areas associated with the field of cytometry, including current, key methodologies and protocols essential to basic and clinical research. Includes new sections on technology principles. Plastic-comb binding, hardcover also available.

Contents of Volume 64

xii

Contributors

xvii

Preface to the Third Edition

xxi

Preface to the Second Edition

xxv

Preface to the First Edition

xxix

PART I Principles of Cytometry and General Methods

A Brief History of Flow Cytometry and Sorting

Myron R. Melamed

Introduction

(1)

Instrumentation

(5)

Applications

(10)

References

(6)

Principles of Flow Cytometry: An Overview

Alice L. Givan

Introduction

(1)

The Illumination of a Particle

(6)

Fluidics: Centering Particles in the Illuminating Beam

(7)

Collection of Light Signals from Particles

(4)

From Light Signals to a Data File

(4)

From Data to Information

(3)

Sorting

(4)

Conclusions

(3)

References

(3)

Laser Scanning Cytometry

Louis A. Kamentsky

Introduction

(1)

Background

(3)

Description of the Instrument

(7)

The Utility and Operational Characteristics of Some Laser Scanning Cytometry List Mode Features

(18)

Utility of Solid Phase Cytometry for Cell Preparation

(2)

Future Directions

(7)

References

(5)

Principles of Confocal Microscopy

J. Paul Robinson

Brief History of Microscope Development

(1)

Development of Confocal Microscopy

(2)

Image Formation in Confocal Microscopy

(4)

Useful Fluorescent Probes for Confocal Microscopy

(4)

Applications of Confocal Microscopy

100

(5)

Conclusions

105

(3)

References

105

(3)

Optical Measurements in Cytometry: Light Scattering, Extinction, Absorption, and Fluorescence

Howard M. Shapiro

Introduction

108

(1)

Signal Processing Tasks in Flow Cytometry: An Overview

108

(2)

The Optical Signal: Interaction of Light with Cells

110

(8)

Detection: Converting Optical Signals to Current

118

(5)

Electronics: Converting Current to Voltage

123

(1)

Fluorescence Compensation and Logarithmic Amplification

124

(3)

Peak Detection, Integration, and Pulse Width Measurement; Triggering

127

(1)

Measurement Sensitivity: Changing Concepts and the Bottom Line

127

(4)

References

129

(2)

Flow Cytometric Fluorescence Lifetime Measurements

Harry A. Crissman

John A. Steinkamp

Introduction

131

(3)

Applications of the Technology

134

(3)

Cell Preparation and Staining

137

(1)

Fluorescence Lifetime Flow Cytometry Instrumentation

138

(3)

Results

141

(3)

Critical Aspects of the Technology

144

(2)

Future Directions

146

(3)

References

147

(2)

Principles of Data Acquisition and Display

Howard M. Shapiro

Introduction

149

(1)

Pulse Characterization Using Analog and Hybrid Circuits

150

(4)

Analog Data Display

154

(1)

Analog-to-Digital Conversion

155

(4)

Pulse Characterization by Digital Signal Processing

159

(2)

Data Storage and Display with Digital Computers

161

(8)

References

167

(2)

Time as a Flow Cytometric Parameter

Larry Seamer

Larry A. Sklar

Introduction

169

(2)

Historical Overview

171

(1)

Sample Mixing and Delivery

171

(4)

Data Analysis

175

(4)

Applications

179

(2)

Conclusions

181

(4)

References

181

(4)

Protein Labeling with Fluorescent Probes

Kevin L. Holmes

Larry M. Lantz

Introduction

185

(1)

Labeling of Proteins with Organic Fluorescent Dyes

186

(8)

Labeling of Proteins with Phycobiliproteins

194

(8)

Conclusion

202

(5)

References

202

(5)

PART II Cell Preparation

Preparation of Cells from Blood

J. Philip McCoy, Jr.

Introduction

207

(1)

Collection, Transport, and Storage of Blood

208

(2)

Fixation and Preservation

210

(1)

Separation of Erythrocytes from Leukocytes

210

(1)

Assessment of Cell Viability

211

(1)

Staining

212

(1)

Summary

212

(6)

Appendix 1

213

(1)

Appendix 2

214

(1)

Appendix 3

214

(1)

References

214

(4)

Cell Preparation for the Identification of Leukocytes

Carleton C. Stewart

Sigrid J. Stewart

Introduction

218

(1)

Antibodies

219

(15)

Tandem Fluorochromes

234

(3)

Cell Preparation and Staining Procedures

237

(7)

Titering Antibodies

244

(5)

Solutions and Reagents

249

(4)

References

250

(3)

Strategies for Cell Permeabilization and Fixation in Detecting Surface and Intracellular Antigens

Steven K. Koester

Wade E. Bolton

Introduction

253

(1)

Application

254

(8)

Materials and Methods

262

(4)

Concluding Remarks

266

(5)

References

266

(5)

PART III Standardization, Quality Assurance

Stoichiometry of Immunocytochemical Staining Reactions

James W. Jacobberger

Introduction

271

(2)

Structure of Immunoglobulin G

273

(1)

Cell Structure

274

(1)

Permeabilized Cell Structure

274

(7)

Antibody-Antigen Reactions

281

(11)

Multiparametric Analyses

292

(2)

Summary

294

(6)

References

295

(5)

Standardization and Quantitation in Flow Cytometry

Robert A. Hoffman

Introduction

300

(1)

General Issues

300

(2)

Performance Characteristics---Dynamic Range, Linearity, Resolution, and Sensitivity

302

(10)

Standardization and Calibration of Common Cytometry Measurements

312

(18)

Examples of Applications Using Calibrated Measurements

330

(2)

Issues in Quantitation of Fluorochromes and Other Molecules

332

(11)

References

334

(9)

PART IV Cell Proliferation

Methods to Identify Mitotic Cells by Flow Cytometry

Gloria Juan

Frank Traganos

Zbigniew Darzynkiewicz

Introduction

343

(2)

Materials

345

(1)

Cell Preparation and Staining

345

(1)

Instruments

346

(1)

Critical Aspects of the Procedure

346

(1)

Results and Discussion

347

(3)

Comparison of Anti-H3-P Monoclonal Antibody with Other Markers of Mitotic Cells

350

(5)

References

353

(2)

Cell Cycle Kinetics Estimated by Analysis of Bromodeoxyuridine Incorporation

Nicholas H. A. Terry

R. Allen White

Introduction

355

(2)

Applications

357

(1)

Materials

358

(2)

Methods

360

(9)

Critical Aspects of the Procedure

369

(7)

References

372

(4)

Flow Cytometric Analysis of Cell Division History Using Dilution of Carboxyfluorescein Diacetate Succinimidyl Ester, a Stably Integrated Fluorescent Probe

A. Bruce Lyons

Jhagvaral Hasbold

Philip D. Hodgkin

Introduction and Background

376

(2)

Reagents and Solutions

378

(1)

Preparation and Labeling of Cells

378

(3)

Gathering of Information Concurrent with Division

381

(5)

Analysis of Data

386

(4)

Application of Carboxyfluorescein Diacetate Succinimidyl Ester to in Vitro Culture of Lymphocytes

390

(2)

Monitoring Lymphocyte Responses in Vivo

392

(3)

Antigen Receptor Transgenic Models

395

(4)

References

397

(2)

Antibodies against the Ki-67 Protein: Assessment of the Growth Fraction and Tools for Cell Cycle Analysis

Elmar Endl

Christiane Hollmann

Johannes Gerdes

Introduction

399

(2)

Application

401

(1)

Materials and Methods

402

(6)

Critical Aspects

408

(1)

Controls and Standards

409

(1)

Examples of Results

410

(9)

References

414

(5)

Detection of Proliferating Cell Nuclear Antigen

Jorgen K. Larsen

Goran Landberg

Goran Roos

Introduction

419

(1)

Molecular Biology of Proliferating Cell Nuclear Antigen

420

(1)

Methods for Immunochemical Detection and Quantification of Proliferating Cell Nuclear Antigen

421

(2)

Results of Cytometric Analysis of Proliferating Cell Nuclear Antigen Expression

423

(3)

Applications in Toxicology, Pathology, and Oncology

426

(7)

References

428

(5)

Lymphocyte Activation Associated Antigens

Andrea Fattorossi

Alessandra Battaglia

Cristiano Ferlini

Introduction

433

(4)

Methodological Aspects

437

(10)

To Flow or Not to Flow for Assessing Lymphocyte Activation/Proliferation? And, If Yes, How Reliable Is Immunophenotyping?

447

(6)

Additional Approaches

453

(2)

Concluding Remarks

455

(12)

References

457

(10)

PART V Cell Death/Apoptosis

Analysis of Mitochondria during Cell Death

Andrea Cossarizza

Stefano Salvioli

Introduction

467

(1)

Scientific Background

468

(2)

Apoptosis and Mitochondria

470

(2)

Method

472

(1)

Results

473

(3)

Pitfalls and Misinterpretation of the Data

476

(1)

Comparison with Other Methods

477

(1)

Reviews of the Applications

478

(1)

Biological and Biomedical Information

479

(1)

Future Directions

480

(7)

References

481

(6)

Cytometry of Caspases

Steven K. Koester

Wade E. Bolton

Introduction

487

(3)

Materials and Methods: Caspase Peptide Inhibitors and Methods to Monitor Responses

490

(7)

Results and Discussion

497

(8)

References

502

(3)

Analysis of Apoptosis in Plant Cells

Iona E. Weir

Introduction

505

(1)

Apoptosis in Plants

506

(3)

Problems Associated with Analyzing Plant Cells Using Flow Cytometry

509

(3)

Morphological Changes of Plant Cells

512

(8)

Physiological Changes during Apoptosis

520

(4)

Conclusion

524

(3)

References

524

(3)

Difficulties and Pitfalls in Analysis of Apoptosis

Zbigniew Darzynkiewicz

Elzbieta Bedner

Frank Traganos

Introduction

527

(2)

Apoptotic Index May Not Be Correlated with Incidence of Cell Death

529

(2)

Difficulties in Estimating Frequency of Apoptosis by Analysis of DNA Fragmentation

531

(1)

The Lack of Evidence Is Not Evidence for the Lack of Apoptosis

532

(1)

Misclassification of Apoptotic Bodies or Nuclear Fragments as Single Apoptotic Cells

533

(2)

Apoptosis versus Necrosis versus ``Necrotic Stage'' of Apoptosis

535

(2)

Selective Loss of Apoptotic Cells during Sample Preparation

537

(1)

Live Cells Engulfing Apoptotic Bodies Masquerade as Apoptotic Cells

538

(1)

The Problems with Commercial Kits and Reagents

538

(1)

Cell Morphology Is Still the Gold Standard for Identification of Apoptotic Cells

539

(2)

Laser Scanning Cytometry: Have Your Cake and Eat It Too

541

(8)

References

544

(5)

PART VI Cell---Cell, Cell--Environment Interactions

Analysis of Cell Migration

Nicole Dodge Zantek

Michael S. Kinch

Introduction and Application

549

(1)

General Strategies to Measure Cell Migration

550

(11)

References

558

(3)

Three-Dimensional Extracellular Matrix Substrates for Cell Culture

Sherry L. Voytik-Harbin

Introduction

561

(2)

Application

563

(2)

Methods

565

(7)

Application of Intestinal Submucosa as a Three-Dimensional Extracellular Matrix Substrate

572

(4)

Summary

576

(7)

References

578

(5)

Three-Dimensional Imaging of Extracellular Matrix and Extracellular Matrix-Cell Interactions

Sherry L. Voytik-Harbin

Bartlomiej Rajwa

J. Paul Robinson

Introduction

583

(1)

Three-Dimensional Imaging of Extracellular Matrix and Extracellular Matrix-Cell Interactions: Current Techniques and Their Limitations

584

(3)

Three-Dimensional Microscopy of Living Systems: Extracellular Matrix and Extracellular Matrix-Cell Interactions

587

(6)

Summary

593

(6)

References

596

(3)

Cytometric Analysis of Cell Contact and Adhesion

Michael S. Kinch

Introduction and Application

599

(1)

General Strategies to Measure Cell-Cell Adhesions

600

(6)

General Strategies to Measure Cell-Ligand Adhesions

606

(2)

Specificity of Cell Adhesion

608

(1)

Optimization of Experimental Conditions

609

(4)

References

611

(2)

Invadopodia: Unique Methods for Measurement of Extracellular Matrix Degradation in Vitro

Emma T. Bowden

Peter J. Coopman

Susette C. Mueller

Introduction

613

(2)

Invadopodia Activity, a Measurement for Localized Membrane Degradation

615

(4)

Fluorescent Activated Cell Sorting---Phagocytosis, a Measurement for Internalization of Proteolyzed Extracellular Matrix

619

(4)

Protocols

623

(6)

References

626

(3)

Index

629

(16)

Volumes in Series

645


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