Dr. P.G.SubramanianAssistan Professor and Officer incharge Hematopathology

laboratoryTata Memorial Hospital

Diagnostic◦ Thrombocytopenia◦ Bicytopenia◦ Pancytopenia◦ Unexplained organomegaly/ Fever/anemia◦ Suspected acute leukemia

Staging for lymphomas Baseline evaluation –◦ Chronic myeloproliferative disorders◦ Myeloproliferative myelodysplastic disorders◦ Plasma cell dyscrasias

Post treatment evaluation◦ Post induction, Day 10 marrows◦ Clinical suspicion of relapse Unexplained cytopenias Unexplained fever

Cytochemistry –◦ Perls iron stain◦ Myeloperoxidase◦ Sudan black B◦ Non specific esterases – ( α-naphthyl acetate or α-

naphthyl butyrate esterase)

Immunophenotyping by flow cytometry Cytogenetics Molecular testing Culture

Normal bone marrow

Megakarocyte

Increased blasts greater than 20%. Acute leukemia/ CML blast crisis (presence of

a documented chronic phase)/ rarely high grade non hodgkins lymphoma

Various morphology of blasts Cytochemistry/ Immunophenotyping

/Cytogenetics/ molecular studies

Myeloperoxidase- If positive Acute Myeloid Leukemia. If negative then immunophenotyping is essential.

Nonspecific esterase – positive in Acute Monoblastic and Acute Monocytic leukemias

Essential for diagnosis of ◦ Acute Lymphoblastic Leukemia (B cell and T cell

type)◦ Acute Myeloid Leukemia M0◦ Acute Monoblastic Leukemia M5◦ Acute Megakaryoblastic Leukemia M7

2 techniques Immunohistochemistry Flow cytometry using

immunofluorescence

Comparison of the two methods currently

Flow cytometry(Immunofluorescence)

Immunohistochemistry

Multiparametric evaluation of single cells

Higher sensitivity

Requires significant concentration of population of interest in the sample

Gives accurate counts and percentages

Fixation and antigen retrieval problems

Morphological confirmation of population of interest

Can detect a few cells in a section

Certain degree of subjectivity in interpretation of weak positivity

Essential for diagnosis of ◦ Philedelphia chromosome positive leukemias and

CML blast crisis◦ Acute promyelocytic leukemia ◦ AML

AML-M3

Case: APML

24/06/2010

8 yr male child referred from outside as case of Acute Lymphoblastic Leukemia.

ThrombocytopeniaBone marrow done – dilute with scattered 40% blasts

like cells and 60% lymphocytesMPO - Negative

FCM - Outside diagnosed as CALLA positive ALL with CD19+, CD10+, HLADR+ & small percentage

of CD34+ cells

R1

Hematogones

Hematogones

Another case : new case4 yrs female child,

High WBC count, thrombocytopenia and low HbBM – 90% blasts, MPO – negative

Immunophenotyping done

R1

HEMATOGONES CALLA+ ALL BLASTS

12 yr old boy with hepatosplenomegaly and lymphadenopathy with complaints of weakness and difficulty in breathing.

Morphology

Diagnosis:T- cell Acute Lymphoblastic Leukemia

5 year old Male child Presented with weakness and severe Pallor. Hb – 4 gm / dL

Given one unit of blood at the local clinicCBC showed few blasts and referred.

Bone marrow on examination shows 45% blasts mixed with lymphocytes and normal myeloid series cells

Diagnosis????

Dyserythropoiesis MDS

43 Yr Male

Hairy Cell Leukemia

HCL

Reed Stern berg Cell