- Email: [email protected]
Clinical flow cytometry: Principles and application
HUMAN
PATHOLOGY
Volume 25, No. 1 (January 1994)
continues to be a puzzle. The suggestion by Drs Min and Seo that multipotential stromal cells might be involved is as feasible as any of the hypotheses that have been advanced so far. NELSON
G. ORD~)P;IEZ,
ADEL
EL-NAGGAR,
K
MD MD
JAE Y. Ro, MD ELVIO
G. SILVA, MD
BRUCE
MACKAY, MD
M. D. Anderson Cancer Center Houston, TX
I.
Wrbster’s New Collrgiate Dictiwwy. Spring&Id, MA, Merriam, 1973 Schmidt D. Dickersin GR, Vawter FG, et al: Wilms’ rmnor: Rrview of ultrastructure and histogenesis. Pathobiol Annu 12:281-300. I982 3. Beckwith JB: Wilms’ twnOr and other renal twmxs of childhood: A selective review from thr National Wilms’ Tumor Studv Patholom. Center. HUM PArHoL 14:481-492.1983 4. Asano T. Fukuda Y, Fukunaga Y, et al: Intra-abdominal dcsmoplastic small cell tumor in an adolescent suggesting a neurogenic origin. Acta Pathol Jpn 43:275-282, 1993 5. Young RH, Eichhorn JH, Dickersin GR, et al: Ovarian involvement by the inua-abdominal desmoplastic small round cell tumw with divergent differentiation: A report of three cases. HUM PATHOL 23:454464, 1992 6. Bian Y, Jordan AC, Rupp M, et al: Effusion cytology of desmoplastic small round cell twmr of the pleura. A case report. Acta Cytol 37:77-82, I993
2.
BOOK REVIEWS Flow Cytometry: Fit
Principles. Alice Longobardi New York, NY, Wiley-Liss, 1992, 202 pages, $34.95.
Givan.
clinical Applications of Flow Cytometry. Roger S. Riley, Edwin J. Mahin, William Ross. New York, NY, Igaku-Shoin, 1993,914 pages, $125.99. Clinical Flow Cytometry: Principles and Application. Kenneth D. Bauer, Ricardo E. Duque, T. Vincent Shankey (eds). Baltimore, MD, Williams 8c Wilkins, 1993, 635 pages, $125. The use of flow cytometry in the practice of anatomic and clinical pathology has expanded considerably in the last decade to provide not only diagnostic information but also useful prognostic data. Instrumentation has improved considerably and relatively simple, “closed” systems can be used for a wide variety of analyses. As the number of available monoclonal antibodies adapted for exploring cell surface markers present on cell suspensions has increased, classification of tumors and other malignancies has demonstrated a parallel increase in complexity. Flow cytometric analysis of nuclear material for DNA ploidy also has seen improvements in standardization of quality control materials and better precision of cell cycle kinetics. However, the, rapid development of these instruments and reagents has outpaced the availability of relevant textbooks for the pathologist with inadequate experience with these techniques, In addition, identifying useful flow information to supplement diagnostic reports has been somewhat haphazard due to a lack of standardization of methods and data interpretation. These three recently released works may help to fill this void. Newcomers to the broad field of flow cytometry may find Dr Givan’s Flow Cytomtq: First Principles to be both informative and enjoyable. The author provides a brief history from the development of the first single cell analyzers to the first fluorescent-detection cytometers using argon laser light sources. The book contains 10 chapters with a strong emphasis on instrumentation. Chapters devoted to application uses of flow cytometry include discussions of lymphocyte analysis (as a pre totype for the quantification of surface markers and techniques of gating) , DNA analysis and molecular biology techniques, and flow cytometry in the clinical laboratory environment, as well as speculation on the future develop ments of the techniques and applications of flow cytometry. The chapters are concise and the general principles are well illustrated. In addition, although the text is not referenced,
110
suggested materials for further reading can be found at the end of each chapter. Figures are used liberally throughout the book to illustrate histograms and instrument configurations. Tables containing data on reagents, absorbance, and CD antigens provide handy reference material. A short glossary helps the novice deal with both the technical vocabulary and the “jargon” that comprise the official language of flow cytometly. This book would be particularly useful to medical technologists with some experience in more traditional a.+ pects of clinical hematology who are involved in setting up a flow cytometlysection or to anyone interested in gaining some basic exposure to the principles of these techniques. Individuals with some familiarity with flow cytometry as a research or diagnostic tool may find this book lacking in specific references and applications. Laboratorians with some exposure to and experience with flow cytometric technology may find Clinical Applications ofFlow Cytomehy by Riley et al to be a more comprehensive text on the subject. Most of the 16 chapters are written (or cowritten) by the three authors, and the first third of the book deals with the basic principles and techniques of flow cytc+ meter operation. Considerable detail is given to the use of control protocols and trouble-shooting artefacts. Specimen preparation and assessment of cell purity are discussed in adequate detail for both surface and DNA techniques. In general, these background chapters are well referenced; there are many recent citations. The clinical application section deals mainly with more established uses of flow cytometry; however, the chapter “Clinical Applications of Immunophenotypic Analysis” is a well-balanced overview of the major diseases in which flow analysis, both ploidy and surface marker studies, has been of particular help in diagnosis and prognosis. A systematic review of the application of flow analysis of various tumors by organ and cell type is presented; however, detailed clinical discussions of some of the more controversial topics in this field are not addressed. Overall, this work is well referenced and provides a detailed background on the techniques of flow cytometry. The goal of the book as stated in the preface is “to serve as a practical source of information for individuals performing flow cytometry, or interpreting flow cytometric data in a health care setting.” This goal certainly has been achieved. The text edited by Bauer et al, ClinicalFlow Cytometry: Bincijhs and ApPlication, is a multi-authored effort intended to “stimulate the intellect of the reader interested in flow cytometry, to provide some insight into the current and emerging
BOOK REVIEWS
clinical applications, and to provide a critical appraisal of its clinical utility.” The 60 or so authors, associate editors, and reviewers have helped ensure the overall high scientific and clinical quality of this text. The 36 chapters cover the basic principles, both theoretical and technical, of flow cytometric analyses as well as applications in hematology and immunology. What sets this work apart is the detailed presentations in clinical oncology. The chapter on lymphomas by Braylan is particularly excellent, and several chapters have an accompanying clinical commentary that openly discusses the advances flow cytometry has contributed to diagnosis and therapeutic decision making as well as the shortcomings and limitations of these techniques. The text is well written and referenced; however, the figures are disappointing in some of the chapters. Pathologists unfamiliar with flow cytometric analysis may find this an invaluable resource in developing a competent background in the clinical use of these techniques. The literature dealing with tumor biology studies using flow analysis has expanded rapidly and any practicing general pathologist will need to be familiar with some of these accepted uses. This text can fill in the voids generated by these advanCeS.-PAUL M. ALLISON, MD, Baylor College of Medicine, Houston,
TX
Tumors and Tumorlike Lesions of the Uterine Corpus and Cervix. P. B. Clement, R. H. Young (eds). New York, NY, Churchill Livingstone, 1993, $95. The goal of the editors of this book (who also wrote most of the chapters) was to concentrate on recent advances in uterine pathology with an emphasis on “newly described entities or. . . significant new information.” They have succeeded. This book is truly comprehensive; although there is a major focus on neoplasms, the editors also present significant insights into the pathology of infectious diseases, notably chlamydial infection. Surgical pathologists will be cheered to know that overall the book places a practical emphasis on routinely stained sections while it also manages to highlight appropriately the occasional problem that is resolved best with additional studies. Some topics are especially timely. An excellent discussion of the resolution of inconsistencies in interpretation of cervical smears and the biopsies that follow will be welcomed by pathologists who take part in departmental quality assurance committees. The place of DNA analysis of HPV lesions is evaluated authoritatively by Christopher Crum in a balanced presentation of pros and cons. Certain old polemics are dealt with in refreshingly practical terms that will be welcomed by pathology residents and seasoned practitioners alike. The difficulties involved in making a distinction between atypical hyperplasia and carcinoma of the endometrium are clearly presented by R.E. Scully and resolved to the extent possible at this time. The photomicrographs are numerous, well selected, and helpful, and the honest conclusion is succinctly stated: “The final diagnosis remains dependent on the interpretation of routinely prepared microscopic sections.” Although trophoblastic tissues are part of the fetus, not the uterus, the problems they cause mostly start in the uterus; their inclusion here is both rational and desirable because it is so useful in the overall context of uterine disease. This chap ter includes an especially useful update on the significant role of DNA ploidy determinations based on the seminal contributions ofJanice Lage, a collaborating author for this volume. In summary, this is a very comprehensive, practical, and useful book, which presents conventional diagnostic surgical pathologic interpretation clearly and authoritatively, while providing an understandable and useful review of the current
implications and applications of molecular techniques those fields with relevant data at this the.-FREDERICK KKAUS, MD, St John’s Merq Medical Center, St Louis, MO
in T.
Fine Needle Aspirationof Palpable Masses. Michael Stanley, Torsten Lowhagen. Boston, MA, Butterworth-Heinemann, 1992, $65. Fine Needle Aspiration of Palpable Masses is the ultimate “how-to-doit” reference on the technical art of the fine needle aspiration (FNA) . While the book appropriately acknowledges the founding contributions of Drs Sixten Franzen and Joseph Zajicek, the authors of this valuable work are themselves outstanding contributors to contemporary FNA. Everyone I know who has had the unique experience of studying under Dr Lowhagen has been impressed by and deeply benefited from Dr Lowhagen’s diagnostic skill, warm rapport with patients, and inspirational teaching style. Through his numerous publications in the field of FNA, Dr Stanley has established his own reputation and now expresses his and Dr Lowhagen’s combined experience in a clear, effective, and concise style. The book is divided into three logical parts. The first two chapters are similar in length, appropriately equating FNA technique with effective patient management and communication skills; these two areas are after all sequentially linked elements of the diagnostic chain. The first chapter, entitled “Equipment, Basic Techniques, Staining Procedures,” methodically and effectively examines issues of needle selection, syringe and syringe holder factors, the actual FNA biopsy sequence, needle motion considerations, and demonstration of the various smearing techniques suitable for particular types of FNA specimens. This chapter concludes with a detailed technical review of appropriate stains and a brief but excellent presentation of highquality color photographs demonstrating selected FNA findings at microscopy. While these few photographs would not comprise an atlas, they show the quality results obtainable with a good technique. The color figures also are an excellent tool for teaching and are an effective means of demonstrating certain microscopic findings to an interested clinician. Indeed, the whole chapter is an excellent educational tool, and on several occasions I have assembled a box of slides, needles, syringes, and a 3inch chunk of liver in a latex glove, and given these along with this book to clinicians I know who could benefit from additional instruction in biopsy technique. The second chapter is entitled “The Patient-Clinical Techniques and Result Reporting.” The novice FNA practitioner, intensely focusing on the details of the FNA biopsy, may not at first appreciate the excellent counsel provided by this chapter. However, after experiencing the anger of a patient or lawyer following an unexpected injury by the FNA biopsy procedure or resulting from a miscommunication on an FNA report, the practitioner will indeed appreciate the depth and experience offered within this second chapter. Here the authors closely examine the various facets of the patient/physician encounter in the FNA clinic. The first section of this chapter begins with “Learning About the Patient’s Problem,” and considers patient positioning depending on the target organ and different approaches for nodules of varying physical characteristics. It concludes with the essential elements of an FNA report as a means of optimizing patient care, balanced by awareness of potential medicolegal issues. The third and final chapter of the book demonstrates the clinical application of the principles outlined in the first chap ters with 10 specific cases from Dr Stanley’s teaching files. These 30 pages provide an appropriate and satisfying ending to a complete and excellent book.
111
PATHOLOGY
Volume 25, No. 1 (January 1994)
continues to be a puzzle. The suggestion by Drs Min and Seo that multipotential stromal cells might be involved is as feasible as any of the hypotheses that have been advanced so far. NELSON
G. ORD~)P;IEZ,
ADEL
EL-NAGGAR,
K
MD MD
JAE Y. Ro, MD ELVIO
G. SILVA, MD
BRUCE
MACKAY, MD
M. D. Anderson Cancer Center Houston, TX
I.
Wrbster’s New Collrgiate Dictiwwy. Spring&Id, MA, Merriam, 1973 Schmidt D. Dickersin GR, Vawter FG, et al: Wilms’ rmnor: Rrview of ultrastructure and histogenesis. Pathobiol Annu 12:281-300. I982 3. Beckwith JB: Wilms’ twnOr and other renal twmxs of childhood: A selective review from thr National Wilms’ Tumor Studv Patholom. Center. HUM PArHoL 14:481-492.1983 4. Asano T. Fukuda Y, Fukunaga Y, et al: Intra-abdominal dcsmoplastic small cell tumor in an adolescent suggesting a neurogenic origin. Acta Pathol Jpn 43:275-282, 1993 5. Young RH, Eichhorn JH, Dickersin GR, et al: Ovarian involvement by the inua-abdominal desmoplastic small round cell tumw with divergent differentiation: A report of three cases. HUM PATHOL 23:454464, 1992 6. Bian Y, Jordan AC, Rupp M, et al: Effusion cytology of desmoplastic small round cell twmr of the pleura. A case report. Acta Cytol 37:77-82, I993
2.
BOOK REVIEWS Flow Cytometry: Fit
Principles. Alice Longobardi New York, NY, Wiley-Liss, 1992, 202 pages, $34.95.
Givan.
clinical Applications of Flow Cytometry. Roger S. Riley, Edwin J. Mahin, William Ross. New York, NY, Igaku-Shoin, 1993,914 pages, $125.99. Clinical Flow Cytometry: Principles and Application. Kenneth D. Bauer, Ricardo E. Duque, T. Vincent Shankey (eds). Baltimore, MD, Williams 8c Wilkins, 1993, 635 pages, $125. The use of flow cytometry in the practice of anatomic and clinical pathology has expanded considerably in the last decade to provide not only diagnostic information but also useful prognostic data. Instrumentation has improved considerably and relatively simple, “closed” systems can be used for a wide variety of analyses. As the number of available monoclonal antibodies adapted for exploring cell surface markers present on cell suspensions has increased, classification of tumors and other malignancies has demonstrated a parallel increase in complexity. Flow cytometric analysis of nuclear material for DNA ploidy also has seen improvements in standardization of quality control materials and better precision of cell cycle kinetics. However, the, rapid development of these instruments and reagents has outpaced the availability of relevant textbooks for the pathologist with inadequate experience with these techniques, In addition, identifying useful flow information to supplement diagnostic reports has been somewhat haphazard due to a lack of standardization of methods and data interpretation. These three recently released works may help to fill this void. Newcomers to the broad field of flow cytometry may find Dr Givan’s Flow Cytomtq: First Principles to be both informative and enjoyable. The author provides a brief history from the development of the first single cell analyzers to the first fluorescent-detection cytometers using argon laser light sources. The book contains 10 chapters with a strong emphasis on instrumentation. Chapters devoted to application uses of flow cytometry include discussions of lymphocyte analysis (as a pre totype for the quantification of surface markers and techniques of gating) , DNA analysis and molecular biology techniques, and flow cytometry in the clinical laboratory environment, as well as speculation on the future develop ments of the techniques and applications of flow cytometry. The chapters are concise and the general principles are well illustrated. In addition, although the text is not referenced,
110
suggested materials for further reading can be found at the end of each chapter. Figures are used liberally throughout the book to illustrate histograms and instrument configurations. Tables containing data on reagents, absorbance, and CD antigens provide handy reference material. A short glossary helps the novice deal with both the technical vocabulary and the “jargon” that comprise the official language of flow cytometly. This book would be particularly useful to medical technologists with some experience in more traditional a.+ pects of clinical hematology who are involved in setting up a flow cytometlysection or to anyone interested in gaining some basic exposure to the principles of these techniques. Individuals with some familiarity with flow cytometry as a research or diagnostic tool may find this book lacking in specific references and applications. Laboratorians with some exposure to and experience with flow cytometric technology may find Clinical Applications ofFlow Cytomehy by Riley et al to be a more comprehensive text on the subject. Most of the 16 chapters are written (or cowritten) by the three authors, and the first third of the book deals with the basic principles and techniques of flow cytc+ meter operation. Considerable detail is given to the use of control protocols and trouble-shooting artefacts. Specimen preparation and assessment of cell purity are discussed in adequate detail for both surface and DNA techniques. In general, these background chapters are well referenced; there are many recent citations. The clinical application section deals mainly with more established uses of flow cytometry; however, the chapter “Clinical Applications of Immunophenotypic Analysis” is a well-balanced overview of the major diseases in which flow analysis, both ploidy and surface marker studies, has been of particular help in diagnosis and prognosis. A systematic review of the application of flow analysis of various tumors by organ and cell type is presented; however, detailed clinical discussions of some of the more controversial topics in this field are not addressed. Overall, this work is well referenced and provides a detailed background on the techniques of flow cytometry. The goal of the book as stated in the preface is “to serve as a practical source of information for individuals performing flow cytometry, or interpreting flow cytometric data in a health care setting.” This goal certainly has been achieved. The text edited by Bauer et al, ClinicalFlow Cytometry: Bincijhs and ApPlication, is a multi-authored effort intended to “stimulate the intellect of the reader interested in flow cytometry, to provide some insight into the current and emerging
BOOK REVIEWS
clinical applications, and to provide a critical appraisal of its clinical utility.” The 60 or so authors, associate editors, and reviewers have helped ensure the overall high scientific and clinical quality of this text. The 36 chapters cover the basic principles, both theoretical and technical, of flow cytometric analyses as well as applications in hematology and immunology. What sets this work apart is the detailed presentations in clinical oncology. The chapter on lymphomas by Braylan is particularly excellent, and several chapters have an accompanying clinical commentary that openly discusses the advances flow cytometry has contributed to diagnosis and therapeutic decision making as well as the shortcomings and limitations of these techniques. The text is well written and referenced; however, the figures are disappointing in some of the chapters. Pathologists unfamiliar with flow cytometric analysis may find this an invaluable resource in developing a competent background in the clinical use of these techniques. The literature dealing with tumor biology studies using flow analysis has expanded rapidly and any practicing general pathologist will need to be familiar with some of these accepted uses. This text can fill in the voids generated by these advanCeS.-PAUL M. ALLISON, MD, Baylor College of Medicine, Houston,
TX
Tumors and Tumorlike Lesions of the Uterine Corpus and Cervix. P. B. Clement, R. H. Young (eds). New York, NY, Churchill Livingstone, 1993, $95. The goal of the editors of this book (who also wrote most of the chapters) was to concentrate on recent advances in uterine pathology with an emphasis on “newly described entities or. . . significant new information.” They have succeeded. This book is truly comprehensive; although there is a major focus on neoplasms, the editors also present significant insights into the pathology of infectious diseases, notably chlamydial infection. Surgical pathologists will be cheered to know that overall the book places a practical emphasis on routinely stained sections while it also manages to highlight appropriately the occasional problem that is resolved best with additional studies. Some topics are especially timely. An excellent discussion of the resolution of inconsistencies in interpretation of cervical smears and the biopsies that follow will be welcomed by pathologists who take part in departmental quality assurance committees. The place of DNA analysis of HPV lesions is evaluated authoritatively by Christopher Crum in a balanced presentation of pros and cons. Certain old polemics are dealt with in refreshingly practical terms that will be welcomed by pathology residents and seasoned practitioners alike. The difficulties involved in making a distinction between atypical hyperplasia and carcinoma of the endometrium are clearly presented by R.E. Scully and resolved to the extent possible at this time. The photomicrographs are numerous, well selected, and helpful, and the honest conclusion is succinctly stated: “The final diagnosis remains dependent on the interpretation of routinely prepared microscopic sections.” Although trophoblastic tissues are part of the fetus, not the uterus, the problems they cause mostly start in the uterus; their inclusion here is both rational and desirable because it is so useful in the overall context of uterine disease. This chap ter includes an especially useful update on the significant role of DNA ploidy determinations based on the seminal contributions ofJanice Lage, a collaborating author for this volume. In summary, this is a very comprehensive, practical, and useful book, which presents conventional diagnostic surgical pathologic interpretation clearly and authoritatively, while providing an understandable and useful review of the current
implications and applications of molecular techniques those fields with relevant data at this the.-FREDERICK KKAUS, MD, St John’s Merq Medical Center, St Louis, MO
in T.
Fine Needle Aspirationof Palpable Masses. Michael Stanley, Torsten Lowhagen. Boston, MA, Butterworth-Heinemann, 1992, $65. Fine Needle Aspiration of Palpable Masses is the ultimate “how-to-doit” reference on the technical art of the fine needle aspiration (FNA) . While the book appropriately acknowledges the founding contributions of Drs Sixten Franzen and Joseph Zajicek, the authors of this valuable work are themselves outstanding contributors to contemporary FNA. Everyone I know who has had the unique experience of studying under Dr Lowhagen has been impressed by and deeply benefited from Dr Lowhagen’s diagnostic skill, warm rapport with patients, and inspirational teaching style. Through his numerous publications in the field of FNA, Dr Stanley has established his own reputation and now expresses his and Dr Lowhagen’s combined experience in a clear, effective, and concise style. The book is divided into three logical parts. The first two chapters are similar in length, appropriately equating FNA technique with effective patient management and communication skills; these two areas are after all sequentially linked elements of the diagnostic chain. The first chapter, entitled “Equipment, Basic Techniques, Staining Procedures,” methodically and effectively examines issues of needle selection, syringe and syringe holder factors, the actual FNA biopsy sequence, needle motion considerations, and demonstration of the various smearing techniques suitable for particular types of FNA specimens. This chapter concludes with a detailed technical review of appropriate stains and a brief but excellent presentation of highquality color photographs demonstrating selected FNA findings at microscopy. While these few photographs would not comprise an atlas, they show the quality results obtainable with a good technique. The color figures also are an excellent tool for teaching and are an effective means of demonstrating certain microscopic findings to an interested clinician. Indeed, the whole chapter is an excellent educational tool, and on several occasions I have assembled a box of slides, needles, syringes, and a 3inch chunk of liver in a latex glove, and given these along with this book to clinicians I know who could benefit from additional instruction in biopsy technique. The second chapter is entitled “The Patient-Clinical Techniques and Result Reporting.” The novice FNA practitioner, intensely focusing on the details of the FNA biopsy, may not at first appreciate the excellent counsel provided by this chapter. However, after experiencing the anger of a patient or lawyer following an unexpected injury by the FNA biopsy procedure or resulting from a miscommunication on an FNA report, the practitioner will indeed appreciate the depth and experience offered within this second chapter. Here the authors closely examine the various facets of the patient/physician encounter in the FNA clinic. The first section of this chapter begins with “Learning About the Patient’s Problem,” and considers patient positioning depending on the target organ and different approaches for nodules of varying physical characteristics. It concludes with the essential elements of an FNA report as a means of optimizing patient care, balanced by awareness of potential medicolegal issues. The third and final chapter of the book demonstrates the clinical application of the principles outlined in the first chap ters with 10 specific cases from Dr Stanley’s teaching files. These 30 pages provide an appropriate and satisfying ending to a complete and excellent book.
111