Table Of ContentFlow Cytometry of Hematological Malignancies
To Angela, my wife and best friend
Flow Cytometry
of Hematological
Malignancies
CLAUDIO ORTOLANI MD
Consultant Clinical Pathologist (retired)
Ospedale dell’Angelo
Venice, Italy
This edition fi rst published 2011, © 2011 by Claudio Ortolani.
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Library of Congress Cataloging-in-Publication Data
Ortolani, Claudio, author.
Flow cytometry of hematological malignancies / Claudio Ortolani, MD, Consultant Clinical Pathologist
(retired), Ospedale dell’Angelo, Venice, Italy.
p. ; cm.
Includes bibliographical references and index.
ISBN 978-1-4443-3588-0 (hardcover : alk. paper) 1. Hematological oncology. 2. Flow
cytometry. I. Title.
[DNLM: 1. Hematologic Neoplasms—diagnosis. 2. Flow Cytometry—methods. WH 525]
RC280.H47O78 2011
616.99(cid:2)418—dc22
2010047400
A catalogue record for this book is available from the British Library.
This book is published in the following electronic formats: ePDF 9781444398045; Wiley Online Library
9781444398069; ePub 9781444398052
Set in 9.25/12pt Minion by MPS Limited, a Macmillan Company, Chennai, India
1 2011
Contents
Foreword by Maryalice Stetler-Stevenson vii CD61 Antigen, 81
CD64 Antigen, 82
Foreword by Bruno Brando ix
CD65 Antigen, 84
Preface and acknowledgments xi CD66c Antigen, 85
CD71 Antigen, 86
Abbreviations xiii
CD79 Antigen, 87
CD103 Antigen, 89
CD117 Antigen, 91
1 Antigens, 1 CD138 Antigen, 92
HLA-DR Antigen, 93
CD1 Antigens, 3 NG2 Antigen, 95
CD2 Antigen, 5 Immunoglobulins, 96
CD3 Antigen, 7 Myeloperoxidase, 100
CD4 Antigen, 15 Cytotoxic Proteins, 101
CD5 Antigen, 19 KIR, CD158 Isoforms, 103
CD7 Antigen, 22 T Cell Receptor, 105
CD8 Antigen, 24 Terminal Deoxy-nucleotidyl Transferase, 108
CD10 Antigen, 27 Bcl-2 Protein, 109
CD11b Antigen, 31 ZAP-70 Protein, 111
CD11c Antigen, 34 References, 112
CD13 Antigen, 36
CD14 Antigen, 40
CD15 Antigen, 42 2 Diseases, 159
CD16 Antigen, 45
CD19 Antigen, 48 Myeloproliferative Neoplasms, 161
CD20 Antigen, 51 Myelodysplastic/Myeloproliferative Neoplasms, 164
CD22 Antigen, 54 Myelodysplastic Syndromes, 165
CD23 Antigen, 56 Acute Myeloid Leukemias, 168
CD24 Antigen, 58 Acute Leukemias of Ambigous Lineage Attribution, 188
CD25 Antigen, 60 Acute Leukemias not Recognized by the WHO
CD30 Antigen, 62 Classifi cation, 191
CD33 Antigen, 63 Neoplastic Diseases of T and B Lymphatic Precursors, 192
CD34 Antigen, 66 Neoplastic Diseases of Mature B Cells, 201
CD38 Antigen, 68 Neoplastic Diseases of Mature T and NK Cells, 234
CD45 Antigen, 70 Appendix, 255
CD45 Isoforms, 75 References, 261
CD56 Antigen, 77
CD57 Antigen, 80 Index, 301
v
Foreword by Maryalice Stetler-Stevenson
Flow cytometry is a crucial tool in the diagnosis of hematol- leukemia or lymphoma. This pattern of organization makes more
ymphoid neoplasms, determining prognosis and monitoring sense than only presenting lists of neoplastic processes and the
response to therapy. Clinical fl ow cytometric immunopheno- expected fl ow cytometric fi ndings. One has to fi rst know the diag-
typing, however, is a complex fi eld requiring extensive expertise nosis on a particular patient before such a reference can be useful.
in normal and abnormal patterns before clinical tests can be Flow Cytometry of Hematological Malignancies also provides the
appropriately interpreted. Those new to the fi eld are left with usual description of typical fl ow cytometric immunophenotypi-
the conundrum of how best to achieve this expertise. At par- cal fi ndings in the various hematolymphoid neoplasms. This is
ticular disadvantage is the resident or clinical fellow seeking to useful as a reference for panel design as well as diagnosis.
interpret fl ow cytometric data on a specifi c patient. The typical Flow Cytometry of Hematological Malignancies is being pub-
fl ow cytometry reference text is written in an encyclopedic for- lished at a time when the fi eld is expanding rapidly and fl ow
mat with extensive narrative that is not conducive to looking cytometry is assuming an even greater role in management of
up the meaning of unusual test results. Furthermore, the gen- patients with hematolymphoid neoplasia. Dr Ortolani, an out-
eral fl ow cytometry textbook, although a useful reference, can- standing fl ow cytometrist, possesses extensive expertise in the
not completely cover all the aspects needed to interpret clinical clinical arena. For over 30 years he was employed in the Clinical
fl ow cytometry data. Therefore, Flow Cytometry of Hematological Pathology Department of the Venice General Hospital, run-
Malignancies fi lls a much needed role in hematopathology and ning one of the fi rst diagnostic fl ow cytometry units in Italy. His
hematology/oncology. The presentation is oriented toward the main clinical activity was the diagnosis of hematological neo-
diagnostic laboratory in the academic center as well as in the plasms, with a particular interest in lymphoproliferative diseases.
general hospital. Dr Ortolani has also been very active in teaching fl ow cytome-
Flow Cytometry of Hematological Malignancies is organized in a try in many national and international courses. He has written
novel manner that makes it especially useful for the medical stu- what I believe to be an outstanding textbook covering the essen-
dent and residents/fellows still in training, while still providing a tial aspects of clinical fl ow cytometry. Dr Ortolani is to be com-
valuable resource for hematopathologists, hematologists/oncolo- mended for this brilliant contribution that is sure to become a
gists and experts in the fi eld of clinical fl ow cytometry. It lists anti- well-used textbook in clinical centers around the world.
gens typically studied in clinical fl ow cytometry laboratories, from
CD1 to CD138, followed by a discussion of general as well as fl ow Maryalice Stetler-Stevenson, PhD, MD
cytometric features and hematolymphoid neoplasms expressing Chief, Flow Cytometry Laboratory
each antigen. Thus, when interpreting a clinical fl ow cytometry National Cancer Institute, National Institutes of Health
report, one can easily research an unusual antigen expressed by a Bethesda, MD, USA
vii
Foreword by Bruno Brando
The European Society for Clinical Cell Analysis (ESCCA) is proud book under the auspices of the ESCCA. It represents a major
to present this volume by Dr Claudio Ortolani. Flow Cytometry achievement for the dissemination of knowledge in one of
of Hematological Malignancies is a benchtop companion for all the most important specialties within clinical cell analysis,
who are involved in the complex process of characterization and as the book aims to improve and standardize the diagnostic
diagnosis of leukemias and lymphomas by immunophenotypical process of malignant blood diseases. As a result, communi-
techniques and fl ow cytometry. cation between clinicians and laboratory operators should
This volume is a useful quick reference text for the matching benefit!
of CD antigens with malignant hematological diseases, as defi ned
by the WHO 2008 classifi cation, taking into account antibody Bruno Brando
clones, features and behavior, with particular emphasis on vari- ESCCA President
ant forms and unexpected presentations. Director, Hematology Laboratory and Transfusion Center,
After several decades of clinical and laboratory practice in Legnano Hospital, Milan, Italy
this field, Dr Claudio Ortolani has meticulously prepared this
ix
Preface and acknowledgments
The cytometric analysis of hematological malignancies is one of The author may have unwittingly sown a number of mis-
the most diffi cult applications of fl ow cytometry, requiring both takes and imprecisions, and he will be grateful to all colleagues
a good knowledge of hematopathology and good control of the who report these to him. He also realizes that this book could
technique. Moreover, the effort of operators is made harder by not have been written without the help of many friends and col-
the continuous evolution of the technology and by the continu- leagues. Being unable to cite all of them, the author wants to par-
ous progress in the comprehension of the nature of the diseases. ticularly thank his friend Bruno Brando, current President of the
This book was compiled from a series of notes originally European Society for Clinical Cell Analysis, for the continuous
intended for people practically involved in the fi eld of diagnostic moral and practical support he has given over the years.
fl ow cytometry, and it is an example of what the author would
have liked to consult at the beginning of his own career. The goal Claudio Ortolani MD
of this book is to offer the reader a quick and updated source of Former Director of the Flow Cytometry Unit
information on the phenotype of the hematological malignan- Clinical Pathology Department
cies recognized by the last WHO classifi cation, with the major Venice General Hospital
exception of Hodgkin lymphoma which because of its peculiar
nature is still beyond the limits of fl ow cytometry, even if things
promise to change in the next few years.
xi
Abbreviations
ABC antibody-binding capacity CINCA chronic infantile neurological cutaneous articular
ABL acute basophilic leukemia syndrome
a-CML atypical chronic myeloid leukemia CLA cutaneous lymphocyte antigen
ADP adenosine diphosphate CM cutaneous mastocytosis
AITL angio-immunoblastic T cell lymphoma CML chronic myeloid leukemia
ALCL anaplastic large cell lymphoma CML-BC CML in blastic crisis
ALCL ALK(cid:3) anaplastic large cell lymphoma, ALK(cid:3) CMML chronic myelomonocytic leukemia
ALK anaplastic lymphoma kinase CMPN chronic myeloproliferative neoplasm
AML acute myeloid leukemia CMV cytomegalovirus
AMLL acute mixed lineage leukemia CNKL chronic natural killer cell lymphocytosis
AMM agnogenic myeloid metaplasia CNL chronic neutrophilic leukemia
ANKL aggressive NK cell leukemia CLPD chronic lymphoproliferative disease
APC antigen-presenting cell CTCL cutaneous T cell lymphoma
ASM aggressive systemic mastocytosis
DFS disease-free survival
ATLL adult T cell leukemia/lymphoma
DLBCL diffuse large B cell lymphoma
ATRA all-trans retinoic acid
AUL acute undifferentiated leukemia EATCL enteropathy-associated T cell lymphoma
AZT azidothymidine (zidovudine) EBV Epstein–Barr virus
EMA epithelial membrane antigen
BAL bronchoalveolar lavage ENKL extranodal NK/T lymphoma
B-ALL B cell acute lymphocytic leukemia ENMZL extranodal marginal zone lymphoma
B-CLL B cell chronic lymphocytic leukemia EPC endothelial progenitor cells
B-CLL/PL B-CLL in prolymphocytoid transformation ET essential thrombocythemia
BCR B cell receptor
BFU-E burst-forming units/erythroid FAB French-American-British
BL Burkitt lymphoma FCCL follicular cell cutaneous lymphoma
B-LBL B lymphoblastic lymphoma FDC follicular dendritic cell
B-NHL B cell non-Hodgkin lymphoma FDCS follicular dendritic cell sarcoma
BPDC blastic plasmacytoid dendritic cell FISH fl uorescence in situ hybridization
B-PLL B cell prolymphocytic leukemia FITC fl uorescein isothiocyanate
B-SLL B cell small lymphocytic lymphoma FL follicular lymphoma
FSC forward scatter
c-ALCL cutaneous anaplastic large T cell
GM-CSF granulocyte macrophage-colony stimulating factor
lymphoma
CEA carcinoembryonic antigen HAL hybrid acute leukemia
CEL chronic eosinophilic leukemia HBLD hairy B cell lymphoproliferative disorder
CFU-GM colony-forming unit granulocyte-macrophage HCL hairy cell leukemia
CGH comparative genomic hybridization HCL-J hairy cell leukemia, Japanese variant
xiii
Abbreviations
HCL-v hairy cell leukemia, variant PAL pyothorax-associated lymphoma
HES hypereosinophilic syndrome PBL plasmablastic lymphoma
HPC hemopoietic precursor cell PCH pseudo Chediak–Higashi
HSTCL hepatosplenic T cell lymphoma PCL plasma cell leukemia
PCMZL primary cutaneous marginal zone lymphoma
ICT indeterminate dendritic cell tumor
PE phycoerythrin
IDCS interdigitating dendritic cell sarcoma
PEL primary effusion lymphoma
Ig immunoglobulin
PHA phytohemagglutinin
IL interleukin
PMA phorbol myristate acetate
ISM indolent systemic mastocytosis
PMF primary myelofi brosis
IVL intravascular lymphoma
PPBL persistent polyclonal B cell lymphocytosis
IVLBCL intravascular large B cell lymphoma
PRCA pure red cell aplasia
JMML juvenile myelomonocytic leukemia PTCL peripheral T cell lymphoma
PTCLnos peripheral T lymphoma not otherwise specifi ed
KIR killer cell immunoglobulin-like receptor
PTLD post-transplant B lymphoproliferative disease
LBCL large B cell lymphoma PV polycythemia vera
LBP lipopolysaccharide-binding protein
RA refractory anemia
LCA leukocyte common antigen
RAEB refractory anemia with excess of blasts
LCH Langerhans histiocytosis
RARS refractory anemia with ringed sideroblasts
LDH lactate dehydrogenase
RCMD refractory cytopenia with multilineage dysplasia
LHL lymphoepithelioid lymphoma
RCUD refractory cytopenia with unilineage dysplasia
LIR leukocyte Ig-like receptors
RN refractory neutropenia
LPL lymphoplasmacytic lymphoma
RT refractory thrombocytopenia
LPS lipopolysaccharide
LRP lung resistance protein s-ALCL systemic anaplastic large cell lymphoma
LyP lymphomatoid papulosis SCF stem cell factor
SLL small lymphocytic lymphoma
MBL monoclonal B cell lymphocytosis
SLVL splenic lymphoma with villous lymphocytes
MCL mantle cell lymphoma/mast cell leukemia
SM systemic mastocytosis
MDCL myeloid dendritic cell leukemia
SM-AHNMD s ystemic mastocytosis with an associated clonal
MDS myelodysplastic syndrome
hematological non-mast cell disorder
MDS-U myelodysplastic syndrome, unclassifi ed
SMZL splenic marginal zone lymphoma
MESF molecules of equivalent soluble
SPTCL subcutaneous panniculitic T cell lymphoma
fl uorochrome
SRCT small round cell tumor
MF mycosis fungoides
SRPL splenic diffuse red pulp small B cell lymphoma
MFI mean fl uorescence intensity
SS Sézary syndrome
MGUS monoclonal gammopathy of undefi ned
SSC side scatter
signifi cance
MHC major histocompatibility complex T-ALL T cell acute lymphocytic leukemia
MM multiple myeloma TAM transient abnormal myelopoiesis
M/NK-AL acute leukemia of myeloid/NK precursors T-CLL T cell chronic lymphocytic leukemia
MoAb monoclonal antibody TCR T cell receptor
MPAL mixed phenotype acute leukemia TCRBCL T cell-rich large B cell lymphoma
MPO myeloperoxidase THL true histiocytic lymphoma
MS myeloid sarcoma T-LBL T cell lymphoblastic lymphoma
MVL microvillous lymphoma T-LGL T cell large granular lymphocytic leukemia
MZL marginal zone lymphoma TMPD transient myeloproliferative disorder
T-PLL T cell prolymphocytic leukemia
NBS Nijmegen breakage syndrome
TPO thrombopoietin
NCA non-specifi c cross-reacting antigen
TZL T zone lymphoma
NCAM neural cell adhesion molecule
NK natural killer WD-EMT well differentiated extramedullary myeloid tumor
NMZL nodal marginal zone lymphoma WHO World Health Organization
xiv
