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Flow cytometry in the diagnosis of mature B‐cell lymphoproliferative

disorders

Article  in  International Journal of Laboratory Hematology · June 2020

DOI: 10.1111/ijlh.13170

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Received: 19 December 2019    Revised: 27 January 2020    Accepted: 31 January 2020

DOI: 10.1111/ijlh.13170

SUPPLEMENT ARTICLE

Flow cytometry in the diagnosis of mature B-cell

lymphoproliferative disorders

Camille Debord  | Soraya Wuillème  | Marion Eveillard  | Olivier Theisen  |

Catherine Godon  | Yanick Le Bris  | Marie C. Béné

Hematology Biology Department, Nantes

University Hospital and CRCINA, Nantes,
France
Correspondence
Marie C. Béné, Hematology Biology, Pôle
Laboratoires, PHU7 9 Quai Moncousu,
44000 Nantes France.
Email: mariecbene@gmail.com
Abstract
B-lineage lymphoproliferative disorders (LPD) are rather frequent diseases, asso-
ciated with specific clinical or biological features but also sometimes of fortuitous
discovery. Multiparameter flow cytometry plays a major role for a rapid diagnostic
indication, on peripheral blood or bone marrow samples in most instances, guiding
complementary analyses and allowing for the proper therapeutic management of
patients. After describing the important pre-analytical precautions required for an
adequate assessment, the immunophenotypic features of small-cell and large-cell
lymphomas are described in this review. The ubiquitous expression of CD19 is a first
mandatory gating step. A possible clonal proliferation is then suspected by the dem-
onstration of surface immunoglobulin light chain restriction. The aberrant presence
of CD5 allows to segregate chronic lymphocytic leukemia and mantle cell lymphoma
in most cases. Other LPD exhibit specific immunophenotypic features. A table of
useful markers and a decision tree are provided. Of note, immunophenotypic data
should as much as possible be interpreted in an integrated manner, involving the
patient's clinical and other biological features, and be completed by further chromo-
somal and/or molecular investigations.

KEYWORDS

B-cell lymphoma, B-cell lymphoproliferative disorders, flow cytometry, immunophenotype,

Matutes score

1 |  I NTRO D U C TI O N
Lymphoproliferative disorders (LPD) are rather common conditions
which can be chronic and indolent or more aggressive diseases.1
Their diagnosis can be prompted by the appearance of enlarged
lymph nodes (LN) or splenomegaly, asthenia, and cytopenia or even
be discovered fortuitously as lymphocytosis on a routine peripheral
blood (PB) complete blood count (CBC). Morphological examina-
tion stands as the first laboratory test suggesting such a condition,
evidencing an increased percentage and/or absolute number and/
or abnormal lymphocytes. This prompts to perform immunopheno-
typic investigations that will reveal, in most cases, the presence of
monotypic B cells. Multiparameter flow cytometry (MFC) thus plays
a critical role in the diagnosis and characterization of B-cell LPDs
with PB and/or bone marrow (BM) involvement. LN biopsies may be
necessary to confirm the diagnosis through pathology investigations
of morphology, immunohistochemistry, or MFC on disaggregated
fresh cell suspensions. Cytogenetics and molecular studies contrib-
ute to a fully integrated diagnosis, useful to guide the proper thera-
peutic strategy.1
This review provides keys to drive the diagnosis of the most
current WHO-defined B-cell LPD by MFC identifying the im-
munophenotypic features of malignant lymphocytes, usually in
PB or BM. This is useful to consider differential diagnoses and to

Int J Lab Hematol. 2020;42(Suppl. 1):113–120. wileyonlinelibrary.com/journal/ijlh |

© 2020 John Wiley & Sons Ltd     113
 |
114       DEBORD et al.
prompt the additional studies mentioned above. Besides general
recommendations, the specific characteristics of the two major
types of LPDs, respectively, of small cell and high grade will be
described.
2 |  PR E-A N A LY TI C A L
R ECO M M E N DATI O N S A N D O R I E NTATI O N
PA N E L
Two major characteristics are fundamental in the investigation of
B-cell LPD, respectively, the detection of monotypy and the expres-
sion of the pan-T antigen CD5.
Monotypy refers to the fact that B cells carry surface immuno-
globulins, as part of the antigen receptor (B-cell receptor or BCR)
complex, necessary for B-cell functions and lymphoma survival. 2 In
a clonal proliferation, all lymphocytes are identical and thus express
the same BCR. Although the demonstration of clonality relies on
molecular studies evidencing the same VDJ sequence of the prolif-
3
erative population, a useful rapid surrogate can be provided by the
demonstration of surface immunoglobulins light chain restriction,
that is, monotypy. Indeed, if a significant population of B cells ex-
press the same light chain, either kappa or lambda, this strongly sug-
gests the presence of an LPD, the normal kappa:lambda ratio being
2:1. Of note, in some rare cases of double LPD, it may happen that
each population expresses a different light chain, mimicking poly-
clonality. Other aberrancies may help to identify such features which
can also be suspected in case of massive B-lymphocyte infiltration.
Moreover, rare cases of monotypic but polyclonal B-cell populations
4
have been reported, for example, in Castleman disease, and vice
versa, some immune reactions may lead to a transient monotypic
expression of light chains by a proliferative clone.
MFC is most often performed on whole PB or BM. Because the
identification of light chain restriction can be impaired by plasma
immunoglobulins, this requires washed PB or BM. Plasma removal
is best performed by using a large volume of isotonic buffer, most
often phosphate-buffered saline, that is, ~50 mL for 100 µL of PB.
After centrifugation for 5  minutes at 600  g, the supernatant is re-
moved and the cell pellet is used to proceed with labeling and red
blood cell lysis. Of note, this will result in some cell loss, without im-
pairing the detection of light chain restriction. However, if a precise
evaluation of the clone size is needed, for instance for the detection
of minimal/measurable residual disease, it is better to use whole PB
in a lysis-no-wash procedure.5
In most cases, the clonal population is predominant and all B
cells express kappa or lambda light chain. Thus, MFC can easily
identify the abnormal population. However, if the clonal population
represents only a part of B cells, it is recommended to use the com-
bination of aberrant markers and light chain expression to better
6
identify the abnormal B-cell population. For this reason, the orien-
tation panel can usefully include CD5, CD20, and CD10 as detailed
below.6 Of course, the presence of CD19 in all combinations is
mandatory to properly gate the whole B-cell population.7 However,
as therapies targeting this surface antigen develop (bispecific anti-
bodies, CAR T cells), care should be taken when tracking LPD cells
after use of such drugs. Of note, this does not apply to the most
current application of MFC in this context, which is diagnosis as
envisioned in this paper.
3 | S M A LL B - C E LL C D5 - P OS ITI V E LPD s
3.1 | Chronic lymphocytic leukemia/small
lymphocytic lymphoma
Chronic lymphocytic leukemia (CLL) is a B-cell neoplasm composed
of monomorphic small mature B cells. It is defined by a PB mono-
typic B-cell count higher than 5 × 109/L. For lower B-cell counts, the
presence of a monotypic population with CLL features character-
izes a monoclonal B lymphocytosis (MBL) or small lymphocytic lym-
phoma.8,9 Of note, in the vast majority of CLL and MBL cases, MFC
examination of a PB sample is sufficient for diagnosis purpose and
there is no reason to test the BM at this stage of evaluation.
Chronic lymphocytic leukemia, often dubbed “the most com-
mon leukemia of adults,” is indeed the most frequently diag-
nosed B-cell LPD, with an annual incidence rate of five cases per
100 000 population.1 Morphologically, CLL cells are small lympho-
cytes, quite fragile, and thus likely to spread on PB smears, yield-
ing ruptured cells called Gümprecht shadows or smudge cells.10
The proliferating population derives from an antigen-experienced
mature CD5+ B cell with mutated or unmutated immunoglobulin
heavy chain.1 MFC on PB is necessary and sufficient to comfort
a diagnosis of CLL. Proliferative B cells present a reduced level
of surface immunoglobulins, which can also be completely ab-
sent in some cases. This translates in the weak expression of one
light chain isotype (kappa or lambda) and of the BCR-associated
complex CD79 (usually tested with anti-CD79b antibodies). CD5,
which is constitutively expressed on T lymphocytes, is positive
in most cases of CLL but with a weaker intensity than on T lym-
phocytes (Figure 1). Another characteristic feature of CLL cells
is the presence of the B-lineage antigen CD23, normally absent
from mature B cells.11 Besides the weak expression of the BCR
complex, CLL cells also downregulate surface CD20 and essen-
tially the CD20 epitope identified by FMC7 which can even be
completely absent. A low fluorescence is also commonly observed
for CD22. The combined assessment of these immunophenotypic
features (CD5, CD22 or CD79b, CD23, FMC7, sIg) provides a sig-
nature initially described by Estella Matutes and thus dubbed
the “Matutes score”12 (Table 1). A Matutes score of 4 or 5 de-
fines CLL. Some atypical CLL cases may score 3 points. In this
case, other markers are useful to help with the diagnosis such as
weak expression of CD81 and upregulated CD43 and, most im-
portantly, CD200, the latter being considered by some as import-
ant as CD23.13 Scores lower than 3 exclude a CLL diagnosis. Rare
DEBORD et al. |
      115

F I G U R E 1   A, Typical chronic
lymphocytic leukemia immunophenotype.
B, Typical mantle cell lymphoma
immunophenotype. The central CD45/
SSC scattergram is gated on leukocytes.
Color code: blue = T cells; magenta = NK
cells; red = normal polytypic B cells;
green = monotypic abnormal cells [Colour
figure can be viewed at wileyonlinelibrary.
com]

TA B L E 1   Immunophenotypic features of the most common B-cell lymphoproliferative disorders

CD5+ CD5− Villous lymphocytes High grade

CLL MCL LPL/WM FL BPLL MZL HCL HCLv SDRPL BL DLBCL

CD19 + + + +/low + + + + + + +
CD5 + + −/+ − − −/+ − − − − −/+
CD23 + low/− −/+ − − −/+ − − − − −/+
sIg low/− + + + ++ + + + + + +/−
CD79b low/− ++ + + ++ ++ + low low low +
CD20 low + + + ++ + ++ + + + +
FMC7 − +/− + + ++ + + + + + +
CD22 low/− + low + ++ + ++ + + + +
CD10 − − − + − − − − − ++ −/+
CD200 ++ − +/− +/− −/+ + ++ + + − −/+
CD43 + −/+ −/+ − + − ++ + − ++ −/+
CD81 low/− + + + + + ++ + + +
CD103 − − − − − − ++ ++ − − −
CD123 − − − − − − ++ − − − −
CD11c −/+ − − −/+ − −/+ ++ + + − −
CD25 −/+ − + −/+ − −/+ ++ − − − +/−
CD13 − − + − − −/+ − − − − −

Note: −/+ = usually negative but may be positive. +/− = usually positive but may be negative. ++ = strong positivity.
Gray cells indicate major discriminating markers.
Abbreviations: BL, Burkitt lymphoma; BPLL, B prolymphocytic leukemia; CLL, chronic lymphocytic leukemia; DLBCL, diffuse large B-cell lymphoma;
FL, follicular lymphoma; HCL, hairy cell leukemia; HCLv, hairy cell leukemia variant; LPL/WM, lymphoplasmacytic lymphoma/Waldenström
macroglobulinemia; MCL, mantle cell lymphoma; MZL, marginal zone lymphoma; SDRPL, splenic diffuse red pulp lymphoma.

CD5-negative CLL cases can only be diagnosed after ruling out
other B-cell neoplasms. 6,14 Of note, CD45 can be more dimly ex-
pressed than on T lymphocytes.
If a CLL diagnosis is confirmed, it can be useful to test for prog-
nostic markers. Investigation of the expression of CD38, CD49d,
or intracytoplasmic ZAP70 has been proposed but is no longer
considered as predictive of a shorter time to treatment.15,16 More
recently, with the development of new drugs for chemo-free ther-
apy, investigation of the mutational status of CLL immunoglobulin
genes has gained a renewed interest since nonmutated heavy chain
genes respond better to BTK inhibitors than to chemotherapy in the
first-line treatment.17
 |
116       DEBORD et al.
3.2 | Monoclonal B-cell lymphocytosis (MBL)
As mentioned above, the diagnosis of MBL is defined by the identi-
fication, by MFC, of a monotypic PB B-cell population representing
<5 × 109 cells/L, without any criterion for another B-cell LPD.8,9 MBL
is classified into three categories based on their immunophenotype,
respectively, (a) CLL-type MBL, (b) atypical CLL-type MBL if CD5 is
expressed with a Matutes score <3, and (c) non-CLL-type MBL. CLL-
type MBL is further classified into two categories, depending on the
9 1,8,9
lymphocyte count being lower or higher than 0.5 × 10 /L. This
is important because, in the first case, there is no evidence that pa-
tients will progress to CLL. CLL-type MBL is reported to be detected
in 3.5%-12% of healthy individuals, and its frequency increases with
1
age. It has been reported that all CLL cases were preceded by a
MBL but, conversely, all MBL will not progress to CLL. Diagnostic
reports should be written carefully because numerous B-cell lym-
phomas can have minimal peripheral dissemination. In case of the
suspicion of atypical CLL-type MBL, it is therefore mandatory to rule
out a leukemic phase of another LPD, especially mantle cell lym-
18
phoma (MCL).
3.3 | Mantle cell lymphoma
Mantle cell lymphoma is a mature B-cell neoplasm deriving from a
peripheral B cell of LN germinal centers' (GC) mantle zone and ac-
1
counts for 3%-10% of non-Hodgkin lymphomas. MCL is usually
considered as an heterogeneous aggressive and incurable disease,
although indolent variants have now been well described.19 At diag-
nosis, BM and sometimes PB can be involved, highlighting the value
of MFC investigations. MCL B cells express CD5 but differ from CLL
by displaying a Matutes score strictly lower than 4, with low or ab-
sent expression of CD23 and moderate to bright expression of CD20
and CD22. CD200 is a very useful marker to differentiate MCL from
CD5+ low-grade LPDs as its expression is extremely rare (<5%) in
MCL 20 (Figure 1B). The rare CD200-positive MCL cases that have
been reported were indolent and non-nodal. In a few cases, MCL B
1
cells can be CD5-negative, yielding an immunophenotype close to
that of other low-grade B-LPDs. Of note, the preferential usage of
lambda light chains by MCL cells has been reported.1 The t(11;14)
translocation, leading to cyclin D1 overexpression, demonstrable by
immunohistochemistry, is typically present in MCL, and cytogenet-
ics must be performed when MFC suggests MCL.
4 |  C D5 - N EG ATI V E B - LPD s
The Matutes score has a limited interest in CD5-negative B-LPDs as
it will always be below 4. A screening tube including CD19, CD20,
immunoglobulin light chains, CD5, and CD10 can be sufficient to ori-
ent the diagnosis in such cases.
4.1 | Lymphoplasmacytic lymphoma and
Waldenström macroglobulinemia
Proliferations of small lymphocytes, plasmacytoid lymphocytes, and
plasma cells, together with a monoclonal peak, constitute a special
entity known as lymphoplasmacytic lymphoma (LPL), provided that
there are no other criteria for other small B-cell lymphomas with
plasmacytic differentiation.1 When these cells infiltrate the BM and
are associated with a plasma peak of IgM (macroglobulinemia), the
disease is named after Jan Waldenström.1 Waldenström macroglob-
ulinemia (WM) is thus typically characterized by splenomegaly and
monoclonal hypergammaglobulinemia of IgM isotype. 21 There are
no specific immunophenotypic features of this disorder. The diag-
nosis relies on the integrated notion of BM involvement, IgM peak,
splenomegaly with or without LN involvement, and morphological
features of lymphoplasmacytoid cells. LPL cells rarely express CD5
or CD10, in <10% of the cases. CD23 is present more frequently
(around 30%).1,22 LPL cells are usually characterized by a low expres-
sion of CD22 and CD25 positivity. 23 Moreover, the presence of a
small contingent of CD13-positive cells has recently been described.
The threshold for significant CD13 expression has thus been set at
1%, based on a large series of B-cell LPDs. 24 WM is the most com-
mon form of LPL. The latter diagnosis is retained on the basis of simi-
lar morphological features of medullary involvement accompanied
by an IgM peak.1 Monoclonal peaks of IgG or IgA isotype can be
observed in rare LPL cases. 25
4.2 | Follicular lymphoma
Follicular lymphoma (FL) is a B-cell neoplasm deriving from sec-
ondary lymphoid organ GC's light zone. 26 FL accounts for about
20% of non-Hodgkin lymphoma and is mostly prevalent in
Western Europe and the United States. It is usually considered
a slowly progressive disease, but BM involvement is observed in
40%-70% of cases. Criteria for progression to diffuse large B-cell
lymphoma (DLBCL) are based on the percentage of large cells
named centroblasts1 assessed in pathology. Of note, FL diagnosis
is mostly based on morphology and MFC cannot help in grading
the disease.
In MFC, FL B cells express the pan-B antigens CD19, CD20, and
CD22, with CD19 often dim. They usually lack CD5 expression but
are characterized by the re-expression of CD10, a marker lost after
the hematogone progenitor stage. 27 Combined with the presence
of morphologically characteristic small cleaved centrocytes, CD10
expression (Figure 2A) is then very evocative of FL. CD5 expression
is rare (3%), and CD5/CD10 double-positive cases must be explored
by the search of t(11;14) and t(14;18) in cytogenetics. 28 Rare cases
of CD10-negative FL have been reported. 29 Immunophenotypic
characterization then becomes less obvious. These cases seem to be
more aggressive and similar to DLBCL. 29
DEBORD et al. |
      117

F I G U R E 2   A, Typical follicular
lymphoma immunophenotype. B, Typical
Burkitt lymphoma immunophenotype. The
top left CD45/SSC scattergrams are gated
on leukocytes. Color code: red = normal
polytypic B cells; green = monotypic
abnormal cells [Colour figure can be
viewed at wileyonlinelibrary.com]

F I G U R E 3   Typical hairy cell leukemia

immunophenotype. The central CD45/
SSC scattergram is gated on leukocytes.
Color code: blue = T cells; magenta = NK
cells; red = normal polytypic B cells;
green = monotypic abnormal cells [Colour
figure can be viewed at wileyonlinelibrary.
com]
 |
118       DEBORD et al.
4.3 | Hairy cell leukemia and hairy cell
leukemia variant
Hairy cell leukemia (HCL) is characterized morphologically by protrud-
ing cytoplasmic expansions providing the cells with a typical “hairy”
30
aspect which drove the disorder's denomination. These B cells, typi-
cally displaying monotypy, appear to be closely related to dendritic
cells since they do express CD123 and CD103. They are also charac-
terized by the presence of surface CD25 and CD11c.31 Of note, the
side scatter of these cells can be misleading, suggesting that they are
monocytes (Figure 3). MFC gating must take this into account. More
important, HCL is usually characterized by pancytopenia or monocy-
topenia, the latter sometimes going unnoticed by CBC instruments
mistaking the HCL cells for monocytes. Conversely, variant HCL show
hyperleukocytosis and no monocytopenia. Cells are morphologically
more difficult to identify but are often characterized by the presence of
a prominent nucleolus and display a variant immunophenotype includ-
1
ing the absence of CD25 and CD123. Moreover, CD43, CD81, and
CD200 have been reported as able to discriminate both HCL types.32
4.4 | B-cell prolymphocytic leukemia
B-cell prolymphocytic leukemia (BPLL) is an extremely rare disease
accounting for about 1% of lymphocytic leukemias.1 This B-cell neo-
plasm mostly relies on a morphological definition. It is considered to
be a proliferation of B-cell prolymphocytes, but CLL in progressive
phase (prolymphocytic or Richter transformation)33 and leukemic
phases of MCL must first be excluded. For a consolidated diagnosis,
prolymphocytes must represent >55% of total PB lymphocytes and
a lymphocytosis over 100 × 109/L is usually observed.
There is no specific BPLL immunophenotype, but pan-B anti-
gens such as CD19, CD20, CD22, CD79b, and FMC7 are strongly
expressed, while CD5 and CD23 are usually but not always absent.
Surface immunoglobulins, identified by light chain monotypy, are ex-
34
pressed with strong intensity.
4.5 | Marginal zone lymphoma
Marginal zone lymphoma (MZL) represents about 10% of non-Hodg-
kin lymphomas.1 This denomination covers different histological
definitions of small B-cell lymphoma. A first entity is that of splenic
MZL (SMZL) composed of small lymphocytes originating from the
GC of splenic white pulp and splenic hilar LN. Such cells can also
be present in BM and PB. In the latter, morphology will disclose
villous lymphocytes in 29%-75% of the cases.35 A second entity is
that of extranodal MZL of mucosae-associated lymphoid tissues
(MALT lymphoma). There, neoplastic cells infiltrate extranodal sites
with a preference for the gastrointestinal tract, especially the stom-
ach. About 2%-20% of the patients will also display BM infiltration.
Finally, nodal MZL involve peripheral LN, BM in one third of the
cases, and occasionally PB (10%-24% of the cases).36
By MFC, MZL is an exclusion diagnosis, that is, after all other
possibilities have been ruled out. Neoplastic cells express the pan-B
antigens CD19, CD20, CD22, and CD24. They are usually negative
for CD10 and CD5. Yet, around 20%-25% of MZL can express CD5,
expression of this antigen being described to be associated with a
higher lymphocytosis and diffuse BM infiltration without impact on
outcome.37 In such cases, exploration of CD200 expression can be
useful to exclude an MCL.
4.6 | Splenic diffuse red pulp small B-cell lymphoma
Splenic diffuse red pulp lymphoma (SDRPL), as well as SMZL, was
originally defined as splenic lymphoma with villous lymphocytes in
the 1989 French-American-British classification.38 Such lymphomas
were described as affecting the splenic white pulp with or without
clinically important red pulp infiltration. In both entities, circulating
lymphocytes, when PB is involved, display short villi, often localized
to one pole of the cell. SDRPL was later described as a splenic lym-
phoma with diffuse involvement of the splenic red pulp and constant
dissemination to the BM and PB. SDRPL remains a WHO provisional
entity that requires further molecular studies to be better charac-
terized.1 However, MFC can contribute to the diagnosis. Neoplastic
SDRPL B cells express strongly CD20 and CD22, moderately CD11c,
and sometimes CD103. CD24, CD25, and CD123 are absent, which
helps to distinguish them from SMZL or HCL. CD5 and CD23 are
positive in about 30% of cases each, yet without coexpression, im-
plying that there is no confusion with CLL.35
5 | H I G H - G R A D E /AG G R E S S I V E LPDS
High-grade lymphomas are aggressive tumors with a high rate of
proliferation. The malignant B cells in such cases are large as shown
in MFC by increased scatter signals.39 The 2016 WHO classification
has clarified this entity and describes numerous types mostly dis-
tinguished by pathological and molecular features.1 MFC is useful in
cases with PB or BM involvement, pleural effusion, ascites, or cer-
ebrospinal fluid (CSF) infiltration. When available, it can complement
pathology on cell suspensions obtained from freshly removed LN.
5.1 | Burkitt lymphoma
Initially described by Dennis Burkitt in African children, this lym-
phoma led to the discovery of the Epstein-Barr virus (EBV) responsi-
ble for these endemic forms. In Western countries, Burkitt lymphoma
is more frequent in children but not necessarily associated with EBV.
It is also observed in immunodeficient patients, especially those with
HIV infection. The gastrointestinal tract is frequently involved (ab-
dominal mass), but central nervous system, PB, and BM infiltration
also occur. In MFC,40 the medium-large basophilic and vacuolated
Burkitt cells express CD45 with a lower intensity than lymphocytes.
DEBORD et al.
F I G U R E 4   Algorithm for the
immunophenotypic diagnosis of B-cell
lymphoproliferative disorders [Colour
figure can be viewed at wileyonlinelibrary.
com]
They are CD19+ CD5- B cells with high expression of CD20 and re-
expression of CD10 (Figure 2B). Monotypic surface immunoglobu-
lins are present (usually IgM) in most cases but may not be detected,
which may lead to the erroneous conclusion of an acute lymphoblas-
tic leukemia. Clinical features and other investigations are crucial in
such cases. This is an emergency diagnosis as treatment should be
initiated promptly. MFC should also be expedited because lympho-
matous cells not only have a high proliferative rate but also are likely
to die quickly. In vivo, this translates into a spontaneous tumor lysis
syndrome contributing to the urgency to initiate management.
5.2 | Diffuse large B-cell lymphoma, NOS
Diffuse large B-cell lymphoma is the most common type of non-
Hodgkin lymphoma which may arise spontaneously or develop as
the transformation of other types of lymphoma (FL, MCL) or CLL
(Richter syndrome), more rarely of MZL or LPL. The diagnosis mostly
relies on pathology and immunohistochemistry. In forms with PB or
BM involvement, MFC can be a useful contribution by identifying
a monotypic population of CD19+ CD20+ cells. CD5 (5%-10% of
cases) and, more often, CD10 (30%-50% of cases) expression can
be observed.40 As for Burkitt lymphoma, the detection of surface
immunoglobulins can be elusive and CD45 expression can be highly
variable but usually near that of lymphocytes. Some authors have
reported a lower expression of CD19 and CD20 in double-hit lym-
phoma, which are high-grade B-cell lymphoma with MYC and/or
BCL2 and/or BCL6 rearrangements, but there is ongoing debate on
the subject.41
6 | CO N C LU S I O N
Multiparameter flow cytometry is a rapid and efficient tool for the
diagnosis of B-cell LPDs in various samples including not only PB and
BM but also effusions, CSF, or disaggregated LN. A comprehensive
set of monoclonal antibodies can be applied for an accurate diagnosis,
|
      119
but a simple orientation panel is useful to guide a rational choice of
markers. This is illustrated in the algorithm presented in Figure 4. As
for most hematological malignancies, knowledge of the morphologi-
cal/cytological features is important to consolidate an integrated di-
agnosis and guide further cytogenetic and molecular investigations.
C O N FL I C T O F I N T E R E S T
The authors have no competing interests.
AU T H O R C O N T R I B U T I O N S
CD and MCB conceived and wrote the review; SW, EM, OT, CG,
and YLB reviewed and edited the manuscript. All authors daily con-
tribute to the diagnosis and follow-up of LPD in Nantes University
Hematology Biology Department.
ORCID
Camille Debord  https://orcid.org/0000-0003-2175-7419
Soraya Wuillème  https://orcid.org/0000-0002-4698-0265
Marion Eveillard  https://orcid.org/0000-0001-7206-5135
Olivier Theisen  https://orcid.org/0000-0003-4578-2627
Catherine Godon  https://orcid.org/0000-0002-0423-0546
Yanick Le Bris  https://orcid.org/0000-0002-0095-6999
Marie C. Béné  https://orcid.org/0000-0002-6569-7414
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How to cite this article: Debord C, Wuillème S, Eveillard M,
et al. Flow cytometry in the diagnosis of mature B-cell
lymphoproliferative disorders. Int J Lab Hematol.
2020;42(Suppl. 1):113–120. https://doi.org/10.1111/
ijlh.13170