77 goolsby wolniakdn3g20un7godm.cloudfront.net/2011/am11fnv/77+flow... · lymphoma cases. the case...

77R (REPEAT) Flow Cytometry: Basic Principles and Case Analysis

Charles Goolsby PhD Kristy Wolniak MD, PhD

2011 Annual Meeting – Las Vegas, NV

AMERICAN SOCIETY FOR CLINICAL PATHOLOGY 33 W. Monroe, Ste. 1600

Chicago, IL 60603

77R (REPEAT) Flow Cytometry: Basic Principles and Case Analysis This session will present basic principles of flow cytometry analysis of hematopoietic malignancies through short lecture and mutlitple case presentations including demonstrated listmode analysis of the cases. Although both acute and chronic processes will be presented, there will be an emphasis on chronic lymphoid and lymphoma cases. The case presentations will also stress the analysis of the flow cytometry results in the context of the morphology and patient presentation.

• Appreication of the variety of flow cytometry analysis approaches necessary for sensitive and specific analysis of hematopoietic malignancies.

• Understand basic technical aspects/prinicples including epitope deletion/masking and defining positive and negative staining including in cases of dim staining.

• Understand the goals and basic guidelines of flow cytometric analysis of hematopoietic malignancies. FACULTY: Charles Goolsby PhD Kristy Wolniak MD, PhD Practicing Pathologists Hematopathology Hematopathology 2.0 CME/CMLE Credits Accreditation Statement: The American Society for Clinical Pathology (ASCP) is accredited by the Accreditation Council for Continuing Medical Education to provide continuing medical education (CME) for physicians. This activity has been planned and implemented in accordance with the Essential Areas and Policies of the Accreditation Council for Continuing Medical Education (ACCME). Credit Designation: The ASCP designates this enduring material for a maximum of 2 AMA PRA Category 1 Credits™. Physicians should only claim credit commensurate with the extent of their participation in the activity. ASCP continuing education activities are accepted by California, Florida, and many other states for relicensure of clinical laboratory personnel. ASCP designates these activities for the indicated number of Continuing Medical Laboratory Education (CMLE) credit hours. ASCP CMLE credit hours are acceptable to meet the continuing education requirements for the ASCP Board of Registry Certification Maintenance Program. All ASCP CMLE programs are conducted at intermediate to advanced levels of learning. Continuing medical education (CME) activities offered by ASCP are acceptable for the American Board of Pathology’s Maintenance of Certification Program.

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Flow cytometry: Basic analysis of hematopoietic malignancies

Charles Goolsby Kristy Wolniak

Northwestern University Medical School

Flow cytometry: Analysis of hematopoietic malignancies

• Immunphenotyping in diagnosis of hematopoietic malignancies– Key, critical component of primary diagnostic

workupworkup– Adjunctive: Critical is correlation with

• Morphology• Other laboratory/pathology data• Clinical data/presentation• Cytogenetics/molecular• History


Lineage determinationB versus TLymphoid versus myeloid

Establishing B cell clonalityBenign (polyclonal) versus malignant (clonal)Clonal implies malignant ???

Diagnostic classificationCLL versus mantle cell lymphoma (MCL)CLL versus mantle cell lymphoma (MCL)Follicular lymphomaHCL

PrognosticZAP-70CD38CD23

Integral component WHO classificationDetection of bone marrow involvement in lymphoma

Sensitive methodology (< 1%)Detection of circulating lymphoma cellsResidual disease detection

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• Potpourri of general analysis comments– +/-– IntensityIntensity– Epitope deletion/masking– Dim staining– General analysis guidelines

• Example case analyses

Analysis of hematopoietic malignancies:What is + and -? Internal Cellular Controls

CD

3 In

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ity

2

3

4

CD

19 In

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ity

Side Scattered Light Intensity0

Side Scattered Light Intensity

CD

3 In

tens

ity

CD7 Intensity

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Analysis of hematopoietic malignancies:Staining Intensity

CD

19

CD

19

CD

19

CD20

DIM

CD20 CD20

MODERATE BRIGHT

Comparison to normal staining intensity

“Quantitative” – number of bound antibodies

Analysis of hematopoietic malignancies:Dim staining comments

• Dim staining = dim antigen• Establish staining level where confident

– Below that, what to do• Competitive inhibition

?

• Competitive inhibition– Epitope not practical, cost prohibitive– Unlabeled antibody, expensive


• Dim staining = dim antigen• Establish staining level where confident

– Below that, what to do• Competitive inhibition

?

p– Epitope not practical, cost prohibitive– Unlabeled antibody, expensive

• Independent antibodies, brighter fluorochrome– No staining not necessarily informative

• Increase or change “blocking” reagents’– Routine 30% FBS– 50% up to 100% NMS– other

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30% FBS 80% NMS

Surface CD3 Surface CD3

CD

45

CD

45

With 50% normal mouse serum With 100% normal mouse serum

Analysis of hematopoietic malignancies:Dim staining

S/N ~ 1.3 S/N ~ 3.1

CD10 CD10

Analysis of hematopoietic malignancies:Epitope deletion/masking

CD

19

CD

19

No staining=no antigen/

-Frequent with Kappa/lambda-Can happen with any antigen

Kappa Lambda

Kappa Lambda

CD

19

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104

105

19

104

105

19

*Independent anti-CD23 antibodies

PE A102 103 104 105

102

103CD

1

CD23*PE A

102 103 104 105

102

103

CD23*

CD

1


nten

sity

nten

sity

CD

3 In

CD7 Intensity*

CD

3 In

CD7 Intensity*

* Independent anti-CD7 antibodies

Analysis of hematopoietic malignanciesGeneral guidelines

• Analysis Goals– Pattern recognition – abnormal patterns

• Subjective, experience needed• Scatter characteristics of a subset

S tt / ti tt• Scatter/antigen pattern• Multiple antigen pattern• Are “unusual”/aberrant patterns abnormal or

reactive change?• Pattern of reactivity with a panel antigens

– Characteristic immunophenotypic signatures• CLL vs mantle cell• Follicular lymphoma/Burkitt’s

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• Account for all cells• Are they normal/abnormal?

• Positive identification of cells of interest• Minimally two + antigens• Minimally one – antigen• More the better• Pan restricted vs associated antigensPan restricted vs associated antigens

• Boolean gating virtually all cases• Scatter gate to remove debris/dead cells, etc• Antigen sets to identify subsets

– Linked to assess other antigen expression• Multiple strategies will be needed• Always assess un-gated for all antigens

– FS vs SS, ……– In way best attuned to note aberrant patterns

• Color eventing/gating• Powerful if done to add information content


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Side Scatter

Side Scatter

Side Scatter

Side Scatter

Side Scatter


atte

rFo

rwar

d S

ca

CD

5

CD

5

Side Scatter CD19CD19

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2

3

4

Sid

e S

catte

red

Ligh

t

CD

19 In

tens

ity

Kap

pa In

tens

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CD

19 In

tens

ity

0 2 6 12 68 10240

Side Scattered Light Forward Scattered Light Lambda Intensity

CD25 Intensity CD103 Intensity CD11c Intensity

CD20 Intensity

CD

19 In

tens

ity

CD

19 In

tens

ity

CD

19 In

tens

ity


CD

19 In

tens

ity

Kap

pa In

tens

ity

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pa In

tens

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Kap

pa In

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CD

19 In

tens

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CD

19 In

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Lambda IntensityKappa IntensityLambda Intensity

Lambda IntensityLambda IntensitySide Scattered Light


CD

19 In

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CD

19 In

tens

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Kap

pa In

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CD

19 In

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Lambda IntensityCD10 Intensity CD5 Intensity

CD20 Intensity CD79b Intensity

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Gate 3

Gate 4

Gate 5

CD

19 In

tens

ity

Kap

pa In

tens

ityCD10 Intensity Lambda Intensity

Kap

pa In

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Kap

pa In

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Kap

pa In

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Lambda IntensityLambda IntensityLambda Intensity

Analysis of hematopoietic malignanciesPatient #1: Case History

• 78 year old female with a relative lymphocytosis

• Asymptomatic

• Laboratory values– WBC 11,300/uL– Hemoglobin 12.9 g/dL– Platelets 195,000/uL

• Peripheral blood

Analysis of hematopoietic malignanciesPatient #1

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CD

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Lymphocytes- Debri

CD3+: 41%

81% 1% 17%

0 256 512 768 10240

0 256 512 768 1024100

100 101 102 103 104100

Side Scatter Side Scatter CD5

100 101 102 103 104100

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100 101 102 103 104100

101

102

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104

CD5 CD16/56/57 CD8

CD

4

CD

5

CD

19

LymphocytesLymphocytes CD3+ cells

CD19+: 55%

NK: 5%

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100

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LymphocytesCD19+ cells

100 101 102 103 104 100 101 102 103 104

100 101 102 103 104100

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Side Scatter CD23Lambda

CD20CD79bFMC7

CD

19

CD

19

CD

19

LymphocytesLymphocytes Lymphocytes

Peripheral blood smear

CD

19

CD5

CD

19

Kap

pa

CD19+ Cells

Analysis of hematopoietic malignanciesMantle cell lymphoma

CD23CD5

CD

19

FMC7

CD

19

CD20

CD

19

CD79b

Lambda

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Analysis of hematopoietic malignanciesMature B cell malignancies-panel of antigens

CD19+/CD19+/ CLLCLL MantleMantle FollicularFollicular HCLHCL@@ MarginalMarginal

CD5 + + - - -

CD10 - - + - -

sIg dim + + + + +

( )CD23 + - - (+) -

CD20 dim + + + +(bright) +

FMC7 - + + + +

CD79b - + + + +

CD11c - (+) - (+) - (+) + -(dim)

CD25 - (+) - - + -

CD103 - - - + -

@ Distinctly increased scatter light intensity

Analysis of hematopoietic malignanciesCLL – variability in archetypical pattern• ~40% of cases “aytpical” (ie moderate or bright)

staining one or more of the pan B cell antigens*– 36% CD20– 19% CD79b

7% FMC7– 7% FMC7• No correlation “atypical” pan B cell antigen

staining intensity and morphology (CLL/PL or transformed) or extent of BM involvement*

• CD23 generally moderate to bright – Can be variable– Subset mantle cell lymphoma CD23+

* Monaghan SA et al, Clinical Cytometry, 2003


CD

19 In

tens

ity

CD

19 In

tens

ity

FMC7 Intensity

CD23 Intensity

Lambda Intensity

CD5 Intensity

Kap

pa In

tens

ity

CD

19 In

tens

ity

Cyclin D1+

t(11;14)+

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25

50

75

100Event-free, CD23-negativeEvent-free CD23-positive

Perc

ent s

urvi

val


• 30-40% CD23+

•28% all sites•11% discordant between sites

• Pertinent clinical differences• Bulky disease (13% vs 39%)

0 10 20 30 40 50 60 70 80 90 100 110 120 1300

25

50

75

100

Overall, CD23-negativeOverall, CD23-positive

Time in Months

Perc

ent s

urvi

val

0 10 20 30 40 50 60 70 80 90 100 110 120 1300

Time in Months

• Splenomegaly (73% vs 42%)

• 4 year EFS: 47% vs 18%, p=0.02

• 4 year OS: 76% vs 52%, p=0.05

• Multivariate Cox regression• CD23, LDH, and HSCT• Hazards ratio (EFS)

• CD23: 0.31, p=0.06• LDH: 0.52, p=0.18• HSCT: 0.99, p=0.99

Keleman et al, AJCP, 2008


• 41 year old male with a rectal mass

• The patient has a history of HIV and recent left lower quadrant pain

• Laboratory values– WBC 1,500/uL (low)– Hemoglobin 8.7 g/dL (low)– Platelets 23,000/uL (low)

• Bone marrow aspirate


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CD19+ cells

0 256 512 768 1024 0 256 512 768 102410

100 101 102 103 10410

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Side Scatter Side Scatter

CD

10

Lambda

CD20

CD19+ cells

PE C 7 A102 103 104 105

102

103

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100 101 102 103 104100

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Lambda Lambda

#1 #2

100 101 102 103 104100

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102

103

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Gate 5

CD19+ cells

#2

#1

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Normal bone marrow

Bone marrow

Bone marrow

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Analysis of hematopoietic malignanciesBurkitt Lymphoma

• Rapidly proliferating B cell lymphoma• Frequently associated with translocation of MYC• Three types of Burkitt lymphoma

– Endemic• Children ages 4-7, predominantly malesChildren ages 4 7, predominantly males• Equatorial Africa• Most are associated with EBV

– Sporadic• Mainly in children and young adults• Low incidence world-wide

– Immunodeficiency-associated• Particularly associated with HIV• Often occurs before CD4+ counts are reduced

• Diagnosis– Clinical features, morphology, immunophenotype, and genetics

• Clinical features– Extranodal sites often involved

• Endemic form – jaw and orbital lesions

– Majority of patients present at an advanced stage

Analysis of hematopoietic malignanciesBurkitt Lymphoma

– Highly aggressive, but can be cured• Characteristic morphology

– Medium-sized cells, basophilic cytoplasm, small round nuclei– Starry sky pattern in solid tissue with admixed tingible body

macrophages– Frequent mitoses

• Genetics– t(8;14)(q24;q32) found in a majority of cases (c-myc and IgH

fusion)

Analysis of hematopoietic malignanciesCD10+ differential

• CD10+, sIg+, clonal/monotypic B cell population

Follic lar– Follicular– Large cell lymphoma– Burkitt’s– Rare immature acute B cell lymphoblastic

leukemia

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• 79 year old female with an incidental finding of lymphadenopathy on imaging

• In good health and asymptomatic

• Laboratory values– WBC 7,000/uL– Hemoglobin 13.2 g/dL– Platelets 230,000/uL

• Lymph node


Forw

ard

Sca

tter

CD

19

CD

19

Side Scatter CD5

Kap

pa

Lambda

CD

19

Kap

pa

LambdaCD10

Side Scatter


CD

20

CD

19

CD

5

CD20

CD79b CD23Side Scatter

CD10 CD10

CD

19

CD

19

CD

19

Lambda

Kap

pa

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• Two monotypic (surface Kappa) CD19+ populations– CD5+, CD10-

• CD23+• CD23+• FMC7-, dim CD79b+• Dimmer CD20+

– CD10+, CD5-• Brighter CD20+• Increased scattered light intensity• FMC7+, CD79b+

Lymph node


• 68 year old male with skin lesions• Multiple erythematous plaque lesions on

thighs and upper extremities. No other symptomssymptoms.

• Laboratory values– WBC 6700/uL – Hemoglobin 14.5 g/dL– Platelets 220,000/uL

• Peripheral blood

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0

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101

102

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CD

3

CD

3

CD

3

CD5 CD2 CD26

CD4 CD8 CD7

Peripheral blood

Peripheral blood

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Mycosis Fungoides/Sezary Syndrome:Pan T cell antigen loss/alteration

• Mycosis Fungoides (MF)– Most common CTCL (~50% of primary cutaneous lymphomas) – Epidermotropic clonal T cell malignancy– Skin lesions (patches, plaques, etc)

• Cells with characteristic “cerebriform” nuclei– Primarily older adults

• Slight male predominance (1 5 to 2:1)• Slight male predominance (1.5 to 2:1)– Indolent

• 85-90% 5 year survival• Sezary syndrome

– Many common pathology/immunophenotypic features with MF– Disease of adults– Skin lesions and generalized lymphadenopathy

• Malignant cells in skin, lymph node, and peripheral blood– Refer to Blood 105:3768, 2005 for diagnostic criteria

– Aggressive disease• ~25% 5 year survival

Analysis of hematopoietic malignanciesMycosis Fungoides/Sezary Syndrome

• Immunophenotype– Mature T cell malignancy (surface CD3+)– Most frequently, T helper immunophenotype

• CD3+CD4+– Characteristically, CD7-

• May be partial– Can be CD7+

• Typically, CD2+, CD5+– But deletion can be seen

– Altered expression of any of the pan T cell antigens can be seen

– Memory cell immunphenotype (CD45RO+, CD29+, etc)– Most frequently, CD25-

• CD25+ in ~10-20% of cases

Analysis of hematopoietic malignanciesCD7 Modulation in Reactive T cells

• Reactive T cells can modulate pan T cell antigen expression– Most temporally short– CD7 can be significant and persistentCD7 can be significant and persistent

• Dim CD7 to CD7- reactive T cells can be CD4+– CD25+, CD26+ (activated immunophenotype)

• Overlaps with MF/Sezary and ATLL immunophenotype– Pattern of CD7 staining can be helpful

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Analysis of hematopoietic malignanciesCD7 Modulation in Reactive T cells

CD7 CD8

CD

3

CD

4


• CD26 T cell activation antigen– Membrane and secreted protein– Modulates chemokine activity– Co-stimulatory molecule (both CD3 and CD2 pathways)Co stimulatory molecule (both CD3 and CD2 pathways)

• Reported to be useful differentiating Sezary cells from reactive T cells– Jones et al, Am J Clin Path 115: 885, 2001– Bernengo et al, Br J Dermatol 144(#1):125, 2001– Circulating Sezary cells frequently CD26-

• Reactive Dim CD7 to CD7- cells which would be CD26+


CD

3

CD

3

CD26CD7

CD7

CD

3

CD26

CD

3

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• How useful?

%

60

80

100

75% 73% 69%

43%

CD4/CD8 >10

>5% convoluted lymphocytes

3

0

20

40

CD26 expression (%)

>1000/mm3 convolutedlymphocytes

Positive TCR generearrangement

• Overall, ~64% of cases show CD4+CD26-• Keleman et al, AJCP, in press, 2007

Analysis of hematopoietic malignanciesLymphoblastic Lymphoma/Leukemia

• 14 year old male• Shortness of breath and cough• Chest discomfortChest discomfort• Chest x-ray revealed a large mediastinal

mass and left pleural effusion• CBC: normal• Received mediastinal mass biopsy


CD

4

CD

4

CD

2

CD

3

CD

5

CD

5

CD8 CD1 CD5

TdT sCD3 cCD3

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Acute Lymphoblastic Leukemia

Bone marrow aspirate Bone marrow core biopsy


• Proliferation of malignant “blast” cells– T-LBL and T-ALL same disease– T-LBL ~80-85% of lymphoblastic lymphomas– Primarily young malesPrimarily, young males

• Most frequently presents with mediastinal mass– Can involve lymph node, spleen, skin, liver– Pleural effusions common– Peripheral blood/bone marrow (if extensive T-

ALL)


• Immunophenotype– Most frequent is “common” thymic T cell

immunophenotype• sCD3-, cCD3+, CD2+, CD5+, CD7+

– Less often dim sCD3 or sCD3+ staining can be seenLess often dim sCD3 or sCD3 staining can be seen• CD4+/CD8+, CD1a+• TdT+, CD34-• Frequently, CD10+• Loss or modulation of CD2, CD5, or CD7 can be seen• Aberrant expression of CD13 and/or CD33 can be seen

– CD79a rarely

– Although rare, more “immature” immunphenotypes can be seen

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• 60 year old male with incidental finding of a mediastinal mass on pre-op evaluation

• Asymptomaticy p

• Laboratory values– WBC 3500/uL – Hemoglobin 14.2 g/dL– Platelets 229,000/uL

Gate 1


Sur

face

CD

3

Forw

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Sca

tter

Cyt

opla

smic

CD

3

CD

7

CD

4C

Sur

face

CD

3

CD5CD5Side Scatter

CD2 CD1a CD8

Mediastinal mass

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Mediastinal massCytokeratin immunohistochemistry


• Frequent differential in patient with mediastinal mass– Thymoma vs T-LBL/T-ALL– Immunophenotype of T-LBL/T-ALL (sCD3-,

cCD3+ CD2+ CD5+ CD7+ CD4+/CD8+cCD3+, CD2+, CD5+, CD7+, CD4+/CD8+, CD1a+, TdT+) can overlap with normal immature thymic T cells

• T cells differentiate in the thymus• Normal thymus mixture of T and non-hematopoietic

cells– Immature, “common thymocyte” T cells– More mature, medullary thymocytes– Mature T cells exiting to the periphery


CD3TdT

CD34

CD2

CD5CD7

CD2

CD5CD3TdT

CD3

CD5

CD2CD3

CD5

CD2

ThymusImmature Peripheral

• Normal is a process of differentiation– Common and medullary populations

• Transition immunphenotypes as well • Pattern of staining can be helpful in differentiating normal vs

abnormal

CD7

CD4/CD8

CD7CD1

CD4 or CD8

CD7

CD4 or CD8

CD7

Common Thymocyte

Medullary Thymocyte

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3

Analysis of hematopoietic malignanciesPattern of expression

sCD

3

CD1a CD8

CD

4

Analysis of hematopoietic malignanciesPattern of expression

Normal BM Archetypical B lymphoblastic leukemia

CD

10

CD20

CD

10

CD20


• 72 year old female with diffuse lymphadenopathy and hepatosplenomegaly

• Anorexia, fatigue, dyspnea, and fevers

• Laboratory values– WBC 13,100/uL (high)– Hemoglobin 8.4 g/dL (low)– Platelets 138,000/uL

• Lymph node

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6

2

8

4

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CD

45

CD

19

22%

77%

0 2 6 12 68 10240

Side Scatter Side Scatter CD5

CD8CD2CD5

CD

3

CD

4

CD

3

T cellsT cells

18%

81%


2

3

4

2

3

4

CD

19

CD

19

0 0

0

2

3

4

0

2

3

4

CD10

CD8CD10

CD20

CD

3

CD

4

All T

0

2

3

4

0

2

3

4

Lymph node

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Lymph node


• 81 y.o. female• Found on floor at home, too weak to

walk/stand (spot easy access to bathroom)• Denies passing out• H/O anemia (refused w/u, takes Fe)• Fatigue 2-3 wks, Myalgias, chills, fever

(100.4F), drenching night sweats • No appetite or regular meals for ~10 days

Analysis of hematopoietic malignanciesPatient #7: Case History cont.

• Treated with antibiotics, mild improvement• Thought she had the flu• Labs

– WBC: 332.1 (~11.0 a few months prior), HGB: 8.2, HCT 25 2 MCV 77 RDW 19 7 PLT 117HCT: 25.2, MCV: 77, RDW: 19.7, PLT:117

• BP 97/52, T 98.6, H 103, RR 16• PE: Alert, “no sign of leukostasis”• Heme/Onc fellow evaluation

– Blood smear: mature appearing lymphocytes– No hemolysis

• Peripheral blood sample received by flow lab

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0 65536 131072 196608 26214

F


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Lymphoblast

Myeloblast

Immature Granulocyte

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CD

45

2 0 2 3 4 5

2

3

4

5

2 0 2 3 4 5

2

2

3

4

5

2 0 2 3 4 5

2

2

3

4

5

CD

2


CD

19

CD

4

CD20CD3 CD7


2

3

4

5

2

3

4

5

CD

45

CD

45

2

3

4

5

2

3

4

5

CD

45

CD

45

CD34

CD13 CD33

CD117


CD

45

MPO

Cyt

opla

smic

CD

3

Cyt

opla

smic

CD

3

CD

19

CD

2

Side Scatter

CD7 CD5

CD79a TdT

CD64

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• ~97% dim CD45, low to intermediate SS intensity cells (<0.1% B cells)– Immature antigens

• CD34+, CD117+• TdT-

Myeloid associated antigens– Myeloid associated antigens• Myeloperoxidase+, CD13+, CD33+• CD64-, CD11b-

– T cell associated antigens• Dim aberrant CD7 and CD5• CD3- (cytoplasmic and surface), CD2-

– B cell associated antigens• CD19-, CD20-, CD79a-

– Other• HLADR+

Myeloperoxidase

Analysis of hematopoietic malignanciesPatient #7: Follow-up

• Follow up– Diagnosis AML– Not treated per family decision– Transferred to Hospice

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Analysis of hematopoietic malignanciesAML

• Flow Cytometric Analysis AML– Lineage – Sub-classification

• No Uitility• Correlations

– Aberrant antigen expression• CD7: ~30 to 35%• CD2: ~20%• CD5: rarer• CD19: ~20%• TdT: ~30%• Helpful in confirming abnormal

Analysis of hematopoietic malignanciesSummary

• Analysis of a panel of antigens– Patterns of staining with multiple antigens– Recognition of normal and abnormal– Account for all cells– + and – antigens for all populations

• Recognition of abnormal (normal) is subjective andRecognition of abnormal (normal) is subjective and pattern recognition driven– Experience needed– +/-, intensity, scattered light characteristics

• Multiple gating/data presentation approaches required• Technical issues including

– Dim staining vs antigen expression– Lack of staining vs lack of antigen (epitope alteration)

• Careful correlation with morphology, pathology, molecular, cytogenetics, ….


• Northwestern Memorial Clinical Flow Cytometry LaboratoryLaura Marzalek Adella KhongJanet McLa ghlin Carolina Ostig inJanet McLaughlin Carolina OstiguinKeisha Hughes Maybelle Tiongson

• Hematopathology TeamLoAnn Peterson Beverly NelsonDiana Variakojis Amy ChadburnYihua Chen Bill Karpus

10/8/2011

30

Flow cytometry: Basic analysis of hematopoietic malignancies

Di i d Q tiDiscussion and Questions

77 goolsby wolniakdn3g20un7godm.cloudfront.net/2011/am11fnv/77+flow... · lymphoma cases. the case...

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