Hematology, Leukocyte Disorders, Instrumentation, and Anemias Review

Flashcards covering hematopoiesis, WBC morphology, maturation stages, automated hematology analyzers, quality control, RBC morphology, and macrocytic/microcytic anemias.

Which mature blood cells develop from the myeloid progenitor lineage?

Erythrocytes, thrombocytes, granulocytes (neutrophils, eosinophils, basophils), and monocytes.

How do Eosin Y and Methylene Blue / Azure B function in Romanowsky staining?

Eosin Y is an acidic/anionic dye with a negative charge that binds basic components to stain eosinophil granules pink-red, cytoplasm pink-red, and RBCs salmon pink. Methylene Blue / Azure B is a basic/cationic dye with a positive charge that binds acidic components (DNA, RNA, basophil granules, nuclear chromatin) to yield blue and purple tones.

What is the normal adult reference interval for total leukocyte (WBC) count?

$$4.8 - 10.8 \times 10^9/\text{L}$$

What are the morphological features, reference interval, and lifespan of mature neutrophils?

They measure $9–15\,\mu\text{m}$, have a multi-lobed nucleus with 2 to 5 lobes connected by thin threads, and contain pale lilac/neutral granules masking clear-blue cytoplasm. Their reference interval is $2.0 - 7.5 \times 10^9/\text{L}$ ($50–70\%$) and their lifespan is 6 hours to a few days.

What are the key morphological features and reference values for eosinophils?

They measure $10–16\,\mu\text{m}$, feature a bi-lobed ('Pince-nez' style) nucleus, are packed with bright orange-red specific granules, have a reference interval of $0.0 - 0.6 \times 10^9/\text{L}$ ($0–4\%$), and circulate for 4 to 5 hours (persisting in tissue for 8 to 12 days).

What are the key morphological characteristics of basophils?

They measure $10–14\,\mu\text{m}$ and feature a bi- or tri-lobed nucleus heavily obscured by large, dark blue/purple water-soluble granules.

What are the key morphological features of a peripheral blood monocyte?

They measure $12–20\,\mu\text{m}$ (or up to $12–22\,\mu\text{m}$), featuring a pleomorphic, indented, folded, or kidney-shaped nucleus (often resembling a 'three-pointed hat') and abundant light blue-gray cytoplasm with a frosted or ground-glass appearance.

What are the six sequential morphological stages of granulocytic maturation?

1. Myeloblast, 2. Promyelocyte, 3. Myelocyte, 4. Metamyelocyte, 5. Band Cell, and 6. Segmented Neutrophil.

How is a promyelocyte morphologically differentiated from a myelocyte?

A promyelocyte ($15–22\,\mu\text{m}$) contains abundant primary azurophilic (non-specific) granules of a burgundy/merlot color, whereas a myelocyte ($10–18\,\mu\text{m}$) marks the initial appearance of secondary (specific) granules alongside a decrease in primary granules and early nuclear flattening.

What morphological criteria define a band cell?

It measures $9–15\,\mu\text{m}$ and has a C- or U-shaped curved nucleus with parallel sides maintained throughout the indentation.

What are the four sequential steps of phagocytosis performed by neutrophils and monocytes?

1. Chemotaxis, 2. Opsonization, 3. Ingestion, and 4. Killing/Digestion.

What is NETosis, and what components make up the extracellular traps?

A process where neutrophils release extracellular traps composed of decondensed DNA, histones, and antimicrobial proteins (such as myeloperoxidase and elastase) to capture extracellular bacteria.

What are the three classic toxic reactive changes observed in neutrophils during severe infection or sepsis?

Toxic granulation (prominent purple-black primary azurophilic granules containing myeloperoxidase), toxic vacuolation (round clear spaces from active phagocytosis and degranulation), and Dohle bodies (light blue-gray oval inclusions of rough endoplasmic reticulum / ribosomal RNA).

What formula is used to calculate the Absolute Neutrophil Count (ANC)?

$$\text{ANC} = \frac{(\%\text{Neutrophils} + \%\text{Bands}) \times \text{Total WBC}\,(10^9/\text{L})}{100}$$

What defines a hematological left shift in peripheral blood?

The presence of immature granulocytes (band cells $> 0.7 \times 10^9/\text{L}$, or the presence of metamyelocytes and myelocytes).

What are the alert and critical reporting threshold values for the Absolute Neutrophil Count (ANC)?

Alert values are ANC $< 1.0 \times 10^9/\text{L}$ or $> 50.0 \times 10^9/\text{L}$. Critical values are ANC $< 0.5 \times 10^9/\text{L}$ or $> 100.0 \times 10^9/\text{L}$.

What protocol must be followed when a 100-cell differential yields zero neutrophils?

Count an additional 100 cells (200 total). If still zero, scan the entire slide before confirming 'rare neutrophil seen' or reporting $0\%$.

What is Pelger-Huët Anomaly (PHA), and how is it distinguished from pseudo-PHA?

An autosomal dominant disorder featuring hyposegmentation where the nucleus forms a symmetrical bilobed ('Pince-nez' / eyeglasses) shape with highly condensed chromatin. True PHA displays $> 70 - 80\%$ bilobed cells, whereas pseudo-PHA (seen in MDS) has fewer.

What are the defining characteristics of May-Hegglin Anomaly?

An autosomal dominant MYH9 disorder characterized by large, hypogranular platelets (causing thrombocytopenia and bleeding) and prominent Dohle bodies in neutrophils and other granulocytes.

How does Alder-Reilly Anomaly present morphologically?

An autosomal recessive condition featuring dense, dark purple-staining cytoplasmic inclusions (Reilly bodies) resembling toxic granulation across neutrophils, monocytes, and lymphocytes, while cell function remains normal.

What clinical and hematological features are seen in Chediak-Higashi Anomaly?

Partial oculocutaneous albinism, photophobia, neutropenia with defective bacterial killing, giant grayish-red lysosomal granules in WBCs, bleeding tendencies, and neurological abnormalities.

What is the clinical purpose of the Leukocyte Alkaline Phosphatase (LAP) score, and which anticoagulant is prohibited?

It differentiates a benign leukemoid reaction (high score) from Chronic Myelogenous Leukemia (CML; low/absent score). EDTA is prohibited because it inhibits the alkaline phosphatase enzyme.

What enzyme deficiency and accumulated metabolite cause Gaucher disease?

Deficiency of $\beta$-Glucocerebrosidase, leading to the accumulation of glucosylceramide.

What enzyme deficiencies cause Niemann-Pick disease and Tay-Sachs disease?

Niemann-Pick disease is caused by a deficiency of Sphingomyelinase (accumulating sphingomyelin). Tay-Sachs disease is caused by a deficiency of Hexosaminidase A (accumulating $\text{GM}_2$-ganglioside).

What are the morphological characteristics of reactive (variant/atypical) lymphocytes?

Enlarged size (up to $15–18\,\mu\text{m}$), intense dark blue peripheral basophilia in the cytoplasm from increased RNA/protein synthesis, and cytoplasm that molds or indents around neighboring red blood cells.

What diagnostic findings and test results characterize Infectious Mononucleosis (IM)?

Caused by Epstein-Barr Virus (EBV / HHV-4), it causes absolute lymphocytosis with abundant reactive lymphocytes (inverted/reverse differential). Diagnosis includes the Monospot latex agglutination test detecting heterophile antibodies (Paul-Bunnell antigen) or specific lateral flow immunoassays.

What are the morphological features of plasma cells, and what do Mott cells contain?

Plasma cells feature an eccentric round nucleus with coarse 'cartwheel' (clock-face) chromatin, deeply basophilic cytoplasm, and a prominent perinuclear hof (Golgi apparatus). Mott cells are plasma cells containing multiple clear globular cytoplasmic inclusions called Russell bodies (accumulated immunoglobulins).

What are smudge cells, and how should they be handled in the laboratory?

Fragile lymphocytes ruptured in vitro during smear preparation, characteristic of Chronic Lymphocytic Leukemia (CLL). If $\ge 10$ smudge cells are seen per 100 WBCs, add them to the total lymphocyte count and comment; they can be prevented by adding 1 drop of $22\%$ bovine albumin to blood prior to slide making.

Which primary hematology analyzer technologies measure cell count/volume versus hemoglobin concentration?

Electrical impedance (Coulter Principle) counts particles (RBCs, platelets) and determines volume from pulse height. Photometry uses a lysing agent and a lamp to measure Hemoglobin (HGB).

What do Forward Scatter, Side Scatter, and Fluorescence represent in flow cytometry?

Forward Scatter (FSC) correlates with cell volume/size, Side Scatter (SSC) correlates with internal complexity and granularity, and Fluorescence (SFL) correlates with the quantity of DNA/RNA.

How do aging commercial QC blood cells affect hematology parameters over time?

HCT, MCV, RDW-SD, PLT-O, and PLT-I tend to increase due to RBC swelling, whereas Monocytes, IGs, Reticulocytes, IRFs, and HPCs tend to decrease.

Why is MCV the preferred parameter for patient delta checks, and what is the primary cause of delta flags?

MCV is the most stable parameter that rarely fluctuates rapidly without a major clinical cause. Pre-analytical human error is the most common reason for delta check flags.

What formula calculates the PAD score in proficiency testing, and what actions do the score ranges require?

$\text{PAD} = \frac{\text{Lab Result} - \text{Assigned Value}}{\text{Allowable Performance Limit}} \times 100$. A score $\le 100\%$ requires no action; $> 100\%$ to $150\%$ triggers a warning (W1); $> 150\%$ triggers mandatory investigative actions and reporting.

What do the three VCS components measure in Beckman Coulter analyzers?

V (Volume) uses direct current impedance for cell size; C (Conductivity) uses high-frequency radio frequency current to penetrate the membrane and assess nuclear structure and internal composition; S (Scatter) uses laser light scatter to evaluate surface structure, shape, and granularity.

What do the four angles of MAPSS technology evaluate in Abbott Cell-Dyn analyzers?

$0^\circ$ (Axial Light Loss) evaluates size/volume; $10^\circ$ evaluates nuclear-to-cytoplasmic complexity; $90^\circ$ polarized side scatter measures cellular granularity; and $90^\circ$ depolarized side scatter measures eosinophilic granules.

What is the clinical significance of dacrocytes, codocytes, and schistocytes?

Dacrocytes (teardrop cells) indicate myelofibrosis, thalassemia, or severe anemias. Codocytes (target cells) indicate excess membrane from liver disease, hemoglobinopathies (Hb C, Hb E), thalassemias, or post-splenectomy. Schistocytes indicate microangiopathic hemolytic anemias (DIC, TTP, HUS, HELLP) or mechanical heart valves.

What are Howell-Jolly bodies, Pappenheimer bodies, and Heinz bodies composed of?

Howell-Jolly bodies are dark purple DNA nuclear remnants. Pappenheimer bodies are clustered iron granules (confirmed with Prussian blue). Heinz bodies are denatured hemoglobin precipitates visible only with supravital stains.

What peripheral blood findings occur following a splenectomy?

Persistent Howell-Jolly bodies, Pappenheimer bodies, target cells (codocytes), leukocytosis, thrombocytosis, and increased poikilocytosis with aging echinocytes.

What is EDTA-induced pseudothrombocytopenia and platelet satellitism?

EDTA-induced pseudothrombocytopenia is in vitro platelet clumping caused by autoantibodies reacting with platelet surface antigens in EDTA blood (corrected with a citrate tube redraw). Platelet satellitism is an in vitro phenomenon where platelets adhere to neutrophil membranes in EDTA.

What are the four main pathophysiological categories of microcytic hypochromic anemias?

1. Disorders of Iron Metabolism (Iron Deficiency Anemia, Anemia of Chronic Inflammation), 2. Disorders of Heme Synthesis (Sideroblastic Anemia), 3. Disorders of Globin Synthesis (Thalassemia syndromes), and 4. Disorders of Porphyrin Synthesis (Lead poisoning).

What role does Hepcidin play in systemic iron regulation?

Hepcidin acts as the gatekeeper hormone: high systemic iron stimulates hepcidin release, which inhibits gastrointestinal iron absorption; low systemic iron suppresses hepcidin to maximize iron absorption.

What is the underlying defect and classic bone marrow finding in Sideroblastic Anemia?

A defect in mitochondrial heme synthesis where iron is trapped inside mitochondria and cannot be incorporated into protoporphyrin. Prussian blue staining reveals ringed sideroblasts encircling erythroblast nuclei in bone marrow.

What are the genetic and hemoglobin profile differences between Beta-Thalassemia Minor and Beta-Thalassemia Major?

Beta-Thalassemia Minor is heterozygous (1 mutated gene on Chromosome 11) presenting with mild microcytic anemia, increased $\text{Hb A}_2$ ($4 - 7\%$), and slightly increased $\text{Hb F}$. Beta-Thalassemia Major (Cooley's Anemia) is homozygous (2 mutated genes) resulting in severe transfusion-dependent anemia, precipitation of excess $\alpha$-chains, hepatosplenomegaly, and secondary hemochromatosis.

What clinical conditions develop when 3 or 4 alpha-globin genes are deleted?

Deletion of 3 genes ($--/-\alpha$) results in Hemoglobin H ($\text{Hb H}$) disease, forming $\beta_4$ tetramers that appear as 'golf ball' inclusions on supravital stain. Deletion of 4 genes ($--/--$) causes Hydrops Fetalis ($\alpha$-Thalassemia Major) producing $\text{Hb Barts}$ ($\gamma_4$), which is fatal in utero.

How does the Mentzer Index differentiate Iron Deficiency Anemia from Thalassemia Minor?

$\text{Mentzer Index} = \frac{\text{MCV}\,(\text{fL})}{\text{RBC count}\,(10^{12}/\text{L})}$. A value $< 13$ indicates Thalassemia Minor, whereas a value $> 13$ indicates Iron Deficiency Anemia.

What peripheral blood smear features distinguish Megaloblastic from Non-Megaloblastic macrocytic anemia?

Megaloblastic anemia exhibits oval macrocytes, hypersegmented neutrophils ($> 5$ lobes), and pancytopenia due to defective DNA synthesis. Non-megaloblastic macrocytic anemia features round macrocytes without hypersegmented neutrophils (commonly associated with liver disease, alcoholism, or hypothyroidism).

What clinical feature distinguishes Vitamin $\text{B}_{12}$ deficiency from pure Folate deficiency?

Vitamin $\text{B}_{12}$ deficiency causes neurological defects (e.g., peripheral neuropathy, dementia, 'brain fog'), which do not occur in pure folate deficiency.

What CBC findings indicate spurious macrocytosis from cold agglutination?

An $\text{MCHC} > 360\,\text{g/L}$ combined with a spuriously low RBC count and a markedly elevated $\text{MCV} > 125\,\text{fL}$.

What are the sequential steps to correct cold agglutination in an EDTA whole blood sample?

1. Verify macroscopic clumping, make a room-temperature smear, and disable auto-release. 2. Incubate the sample in a $37^\circ\text{C}$ water bath for 20 to 30 minutes. 3. Re-mix, prepare a warm smear, re-analyze, and verify MCHC normalizes below $360\,\text{g/L}$. 4. If unresolved, perform warm plasma replacement: centrifuge at $1500\,\text{rpm}$ for 10 minutes, replace plasma with equal volume pre-warmed saline or diluent, incubate 15 to 20 minutes at $37^\circ\text{C}$, and re-assay.