Leukemia Flashcards
Leukemias vs. Lymphomas: Overproduction of cells
Leukemias: Overproduction of various types of immature or mature cells in the bone marrow and/or peripheral blood. Lymphomas: Solid malignant tumors of the lymph nodes and related WBCs in bone tissue.
Leukemias vs. Lymphomas: Cell Types
Leukemias: Frequently involves WBCs of the myelogenous or lymphocytic cell types. Lymphomas: The distinctive cell type is the lymphocyte.
Leukemias vs. Lymphomas: Blood-brain barrier
Leukemias: Malignant cells easily trespass the blood-brain barrier. Lymphomas: Malignant cells are initially confined to lymph nodes, spleen, liver, and bone marrow, though they can spill into the circulating blood.
Leukemias vs. Lymphomas: Tumor location and spread
Leukemias: Malignant cells primarily in bone marrow and peripheral blood. Lymphomas: Solid tumors in lymph nodes and organs containing mononuclear phagocytic cells, may spill over into circulating blood.
Leukemia: Definition
Overproduction of immature or mature WBCs in the bone marrow or peripheral blood.
Leukemia: Disease course and WBC count
More blasts = shorter, fatal course; ↑ WBC with left shift.
Leukemia: M:E ratio
10:1.
Anemia type in acute leukemia
Normocytic, normochromic.
Acute Leukemias: Duration and cell forms
Short duration; numerous immature cells in bone marrow/peripheral blood; ↑ WBC count.
Chronic Leukemias: Duration and cell forms
Long duration; mostly mature cells in bone marrow/peripheral blood; WBC count varies (elevated or lower than normal).
FAB Classification of Leukemias
Based on morphology in Romanowsky-stained smear and cytologic/histochemical characteristics of cells.
Myeloperoxidase (MPO): Function
Used to differentiate AML blasts from ALL blasts.; found in primary granules of neutrophils, eosinophils, and monocytes
MPO Positive Cells
Neutrophilic granulocytes, Auer rods, leukemic blasts in FAB M1, M2, M3, eosinophils.
MPO Weakly Positive or Negative Cells
Monocytes.
MPO Negative Cells
Myeloblasts, basophils, lymphocytic and erythrocytic cell series.
Peroxidase Stain Reminders
May produce red-brown, dark brown, or black color; RBCs may turn brown due to pseudoperoxidase activity in hemoglobin.
Precautions for DAB Handling
Wear protective clothing, use mechanical pipetting aids, clean up spills instantly, wash hands after use, weigh benzidine in hood.
Peroxidase Enzyme Sensitivity
Sensitive to light; stain immediately or keep in dark. Older smears or those exposed to light should not be peroxidase negative.
Cyanide-resistant Peroxidase Stain
Detects eosinophilic leukemia; eosinophil peroxidase differs due to enzyme activity with sodium cyanide.
Sudan Black B (SBB): Function
Stains sterols, neutral fats, phospholipids in primary and secondary granules of neutrophils and lysosomal granules of monocytes.
SBB Positive Cells
Promyelocyte, myelocyte, metamyelocytes, bands, segmented neutrophils (strongly positive), leukemic blasts, Auer rods, eosinophils.
SBB Weakly Positive or Negative Cells
Myeloblasts, monocytic cells.
SBB Negative Cells
Lymphocytes and precursors, megakaryocytes, platelets, erythrocytes.
SBB Staining Reminders
Brownish-black granules in myelocytic precursors, few granules in monocytes, eosinophilic granules with central pallor.
SBB Staining Properties
Can be performed on specimens months old; reagents not carcinogenic.
SBB Disadvantages
Time-consuming (1-2 hours), possible false positives in lipid vacuole disorders, increased background in bone marrow specimens.
Lymphocytic Leukemias: Myeloperoxidase and Sudan Black B
Myeloperoxidase: negative, Sudan Black B: negative.
Most Common Childhood Leukemia
Acute Lymphocytic Leukemia (ALL).
FAB Classification: L1
70% of childhood ALL; markers: CALLA (CD10), TdT, CD19, CD20.
FAB Classification: L2
70% of adult ALL; markers: TdT.
FAB Classification: L3
Rare in children/adults; markers: sIg, CD19, CD20, CD22, CD24.
FAB Classification: Cell Size L1
Homogeneous population of small blasts.
FAB Classification: Cell Size L2
Heterogeneous population of large blasts.
FAB Classification: Cell Size L3
Homogeneous population of large blasts with nuclear and cytoplasmic vacuoles.
Nucleus L1
Uniformly round, small.
Nucleus L2
Irregular.
Nucleus L3
Round to oval.
Nucleolus L1
Single.
Nucleolus L2
Single to several.
Nucleolus L3
Two to five.
Chromatin L1
Slightly reticulated with perinucleolar clumping.
Chromatin L2
Fine.
Chromatin L3
Coarse with clear parachromatin.
Cytoplasm L1
Scant, blue.
Cytoplasm L2
Moderate, pale.
Cytoplasm L3
Moderate, blue, prominently vacuolated.
Cyto-chemistry L1
PAS+: Positive, Methyl Green Pyronin: Negative, ORO: Positive (sometimes).
Cyto-chemistry L2
PAS+: Positive, Methyl Green Pyronin: Negative, ORO: Positive (sometimes).
Cyto-chemistry L3
PAS: Negative, Methyl Green Pyronin: Positive, ORO: Positive.
Immunologic Markers: E Rosettes
T-ALL: Positive.
Immunologic Markers: Surface Ig
B-ALL: Positive.
Immunologic Markers: Serum Anti-ALL
Common ALL: Positive.
Most common type of leukemia in elderly, characterized by persistent lymphocytes
Chronic Lymphocytic Leukemia (CLL).
Chronic Lymphocytic Leukemia (CLL) cells present
Increased smudge cells, Rieder cells in peripheral blood smear.
Clinical Variations of CLL
Hairy-cell leukemia, Lymphosarcoma cell leukemia, Prolymphocytic leukemia.
Acute lymphoblastic leukemia (ALL) Solid tumor counterpart
Lymphoma, poorly differentiated; lymphocytic.
Chronic lymphocytic leukemia (CLL) Solid tumor counterpart
Lymphoma, well-differentiated; lymphocytic.
Monocytic leukemia Solid tumor counterpart
Reticulum cell sarcoma.
Acute myelogenous granulocytic leukemia Solid tumor counterpart
Chloroma.
Plasma cell leukemia Solid tumor counterpart
Myeloma.
Stem cell leukemia Solid tumor counterpart
Lymphoma, undifferentiated.
Non-lymphocytic leukemias/myelogenous leukemias (granulocytes, monocytic progenitor) Initial stains: Myeloperoxidase and Sudan Black reaction
Positive.
Acute Myelogenous Leukemia (AML) M0 - Myelocytic
AML, minimally differentiated/undifferentiated. MPO and SBB negative.
Acute Myelogenous Leukemia (AML) M1 - Myelocytic
AML, without maturation. May demonstrate Auer rods.
Acute Myelogenous Leukemia (AML) M2 - Myelocytic
AML, with maturation. Most common subtype of AML. May demonstrate Auer rods.
Acute Myelogenous Leukemia (AML) M3 - Myelocytic
Acute Promyelocytic Leukemia (APL), associated with DIC and Faggot cells. Demonstrates Auer rods.
Acute Myelogenous Leukemia (AML) M3V - APL, microgranular variant
Cells have characteristic ‘butterfly’, ‘bowtie’, ‘coin-on-coin’, or ‘apple core’ nuclei.
2nd most common subtype of AML. May demonstrate Auer rods.
Acute Myelomonocytic Leukemia (AMML) M4 - Myelocytic Monocytic
AMML with increased marrow eosinophils.
Acute Myelomonocytic Leukemia (AMML) M4E
Schilling Leukemia.
Acute Monocytic Leukemia (AMOL) M5 - Monocytic
AML poorly differentiated. Seen in children; >80% monoblasts in BM.
Acute Monocytic Leukemia (AMOL) M5A
Well differentiated. Seen in middle-aged adults; <80% monoblasts in BM.
Acute Monocytic Leukemia (AMOL) M5B
Di Guglielmo’s Syndrome. Demonstrates Auer rods.
Acute Erythroleukemia M6 - Erythrocytic Myelocytic
AML M6 reaction with PAS. Strongly positive (L1, L2, M6).
Acute Erythroleukemia M6
Di Guglielmo’s Syndrome anemia type: macrocytic, normochromic.
Anemia Type: Di Guglielmo’s Syndrome
Acute Megakaryocytic Leukemia requires immunocytochem staining for accurate diagnosis.
M7 - Megakaryocytic, Factor VIII stains positive.
Acute basophilic leukemia.
AML 8 - Acute Basophilic Leukemia.
M1, M2, M3 leukemia positive cytochemical stains.
Positive for MPO, SBB, Naphthol AS-D, Chloroacetate (SE).
M4 (AMML) leukemia positive cytochemical stains.
Positive for MPO, SBB, Naphthol AS-D, Chloroacetate (SE), α-Naphthyl Butyrate Esterase (NSE), α-Naphthyl Acetate Esterase (NSE).
M5 (AMoL) leukemia cytochemical stains.
Positive or negative for SBB, positive for α-Naphthyl Butyrate Esterase (NSE), α-Naphthyl Acetate Esterase (NSE).
M6 leukemia cytochemical stains.
Positive or negative for MPO, SBB, Naphthol AS-D, Chloroacetate (SE).
M7 leukemia cytochemical stains.
Localized positivity for α-Naphthyl Acetate Esterase (NSE) and positive for Factor VIII stain.
Myelocytic leukemia positive cytochemical stains.
Positive for MPO, SBB, SE, Naphthol AS-D.
Monocytic leukemia positive cytochemical stains.
Positive for NSE, Butyrate Esterase.
ICC Classification AML subtypes with ≥10% blasts
Includes t(15;17)/PML::RARA, t(8;21)/RUNX1::RUNX1T1, inv(16)/CBFB::MYH11, t(9;11)/MLLT3::KMT2A, t(6;9)/DEK::NUP214, t(3;3)/GATA2::MECOM, mutated NPM1.
ICC Classification AML subtypes with ≥20% blasts
t(9;22)/BCR::ABL1, myelodysplasia-related gene mutations, myelodysplasia-related cytogenetic abnormalities.
WHO 2022 Classification usually represents
No blast threshold for classification.
WHO 2022 Classification AML subtypes with ≥20% blasts
BCR::ABL1 fusion, CEBPA mutation, myelodysplasia-related, defined by differentiation.
Chronic Myelogenous Leukemia (CML) AKA,
Chronic granulocytic leukemia
Chromosome Associated with CML,
Philadelphia Chromosome (Ph1)
First description of Philadelphia Chromosome,
Peter C. Nowell, 1960
Cause of Philadelphia Chromosome,
Reciprocal translocation between chromosomes 9 and 22
Result of translocation in CML,
Formation of BCR-ABL1 fusion gene
Three clinical phases of CML,
Chronic phase, Accelerated phase, Blast crisis
Percentage of CML patients with Philadelphia Chromosome,
0.9
Significance of Philadelphia Chromosome in CML,
Indicates good prognosis
Chromosomes involved in CML translocation,
Chromosome 9 and 22
Parts of chromosomes involved in translocation,
Long arms of chromosomes 9 and 22
Must be differentiated from Chronic Myelogenous Leukemia (CML) because it may cause confusion,
LEUKEMOID REACTION (LR)
LEUKEMOID REACTION (LR) is characterized by,
Excessive leukocytic response in peripheral blood
WBC count in LEUKEMOID REACTION (LR),
Greater than 50 X 10⁹ /L (with neutrophilia and marked left shift)
Marked left shift in LEUKEMOID REACTION (LR),
Presence of immature neutrophilic forms
Most frequent WBC increase in LEUKEMOID REACTION (LR),
Neutrophils
Generally used to distinguish LR from CML,
LEUKOCYTE (NEUTROPHIL) ALKALINE PHOSPHATASE (LAP/NAP) TEST
Principle of LAP/NAP Test,
↑ LAP activity observed in neutrophils that have undergone normal growth
KAPLOW’S METHOD Principle,
Hydrolysis of sodium alpha naphthyl phosphate by alkaline phosphatase produces a colored precipitate with a diazotised amine
Fixative used in KAPLOW’S METHOD,
Methanol and formalin
Buffer used in KAPLOW’S METHOD,
Propanediole
Substrate in KAPLOW’S METHOD,
Sodium alpha naphthyl phosphate
Initial stain used in KAPLOW’S METHOD,
Brentamine fast game salt
Counterstain used in KAPLOW’S METHOD,
Aqueous Mayer’s hematoxylin
Procedure for LAP Test (Kaplow’s Method),
Immerse dry blood smear in fixative for 30 seconds, then dry
Working substrate for LAP test,
Made up of buffer, substrate, and initial stain
After applying working substrate in KAPLOW’s LAP,
Allow to stand for at least 10 minutes
Counterstaining time for KAPLOW’s LAP,
10 to 15 minutes
After counterstaining in KAPLOW, where is it mounted,
Glycerol
Examination of KAPLOW’s LAP under the microscope,
Look for reddish-brown to black precipitate in neutrophil cytoplasm
Scoring of neutrophils in LAP test,
Count 100 segmented neutrophils and bands, score based on precipitate
Score for no precipitate,
0
Score for slightly diffused precipitate,
1+
Score for moderately diffused precipitate,
2+
Score for heavily diffused precipitate,
3+
Score for very heavily diffused precipitate,
4+
Normal LAP score range,
15 to 100
Interpretation of normal LAP score,
Leukomoid reaction
Interpretation of decreased LAP score,
CML
Examples of cases with INCREASED KAPLOW’S (LAP) SCORE,
Third trimester pregnancy, PCV, Infections, Intoxication
Examples of cases with DECREASED KAPLOW’S (LAP) SCORE,
CML, Paroxysmal nocturnal hemoglobinuria, Sideroblastic anemia, Myelodysplastic syndrome
Leukocyte types in CML vs. LEUKEMOID REACTION,
CML: Blasts/promyelocytes, LEUKEMOID REACTION: Myelocytes
Toxic granulation in CML vs. LEUKEMOID REACTION,
CML: Absent, LEUKEMOID REACTION: Present
Eosinophils/Basophils in CML vs. LEUKEMOID REACTION,
CML: Decreased (↓), LEUKEMOID REACTION: Increased (↑)
LAP score in CML vs. LEUKEMOID REACTION,
CML: Decreased (↓), LEUKEMOID REACTION: Increased (↑)
Philadelphia chromosome in CML vs. LEUKEMOID REACTION,
CML: Usually present, LEUKEMOID REACTION: Absent
Splenomegaly in CML vs. LEUKEMOID REACTION,
CML: Usually prominent, LEUKEMOID REACTION: Mild (if present)
Platelet count in CML vs. LEUKEMOID REACTION,
CML: >600 or <50 X 10⁹/L, LEUKEMOID REACTION: Normal