Acute Leukemia Diagnostics and Cytochemistry

Flashcards covering cytochemical staining principles, diagnostic utility, FAB vs. WHO classification systems, and advanced immunophenotypic markers for acute leukemias.

What is the biochemical principle of the Myeloperoxidase (MPO) stain?

The intracellular MPO enzyme uses hydrogen peroxide ($H_2O_2$) to catalyze the oxidation of an indicator substrate, such as diaminobenzidine, resulting in an insoluble brown or black-blue precipitate.

How is a blast classified as MPO-positive in diagnostic utility?

A blast is classified as MPO-positive if it demonstrates even a single, well-defined black/brown granule.

What makes Sudan Black B (SBB) useful for identifying myeloblasts in aged specimens?

SBB is a non-enzymatic, lipophilic dye that partitions into structural lipids; unlike the MPO enzyme, these lipid structures remain stable after thermal protein desaturation or specimen aging.

How do Sodium Fluoride (NaF) ions affect Non-Specific Esterase (NSE) in monocytic versus granulocytic lineages?

In monocytic lineages (M4 and M5), NaF causes $100\%$ inhibition of the enzyme; in granulocytic lineages (M1, M2, and M3), any baseline positivity is resistant to NaF inhibition.

Describe the Periodic Acid-Schiff (PAS) staining pattern characteristic of lymphoblasts in Acute Lymphoblastic Leukemia (ALL).

A coarse, highly localized positivity manifesting as large chunky blocks or a "string of pearls" ring surrounding the nucleus, set against a clear cytoplasmic background.

What is the clinical significance of Acid Phosphatase Isoenzyme 5 in Hairy Cell Leukemia?

This specific isoenzyme possesses a unique tertiary molecular architecture that makes it resistant to inhibition by L-tartaric acid, allowing it to maintain an intense orange-red stain in the TRAP assay.

What is an Auer rod and what is its diagnostic implication?

An Auer rod is a crystalline inclusion formed by the fusion of primary azurophilic granules; its presence definitively excludes Acute Lymphoblastic Leukemia (ALL).

How does the blast threshold for an acute leukemia diagnosis differ between the FAB and WHO systems?

The FAB system requires $\ge 30\%$ blasts in the bone marrow or blood, whereas the WHO system reduced the threshold to $\ge 20\%$ (with exceptions for specific genetic abnormalities).

What is the characteristic immunophenotype of Acute Promyelocytic Leukemia (AML M3)?

This subtype is typically $CD34$-negative and $HLA-DR$-negative, while strongly expressing myeloid markers such as $CD13$ and $CD33$.

Why is the FAB M3 variant considered a critical medical emergency?

The presence of hypergranular promyelocytes packed with procoagulant enzymes poses a high risk for the development of Disseminated Intravascular Coagulation (DIC).

In flow cytometry gating, how is a Myeloid lineage assigned for Mixed Phenotype Acute Leukemia (MPAL)?

Myeloid assignment requires a flow index of $\ge 1.5$ for intracellular Myeloperoxidase (iMPO) or the expression of at least two monocytic markers such as $CD14$, $CD64$, $CD11c$, or $CD36$.

What are the three tiers of T-ALL maturation based on surface markers?

1. Pro-T ($cCD3+$, $CD7+$, $sCD3-$, $CD1a-$, $CD4-/CD8-$); 2. Cortical ($cCD3+$, $CD1a+$, $CD4+$, $CD8+$); 3. Medullary ($sCD3+$, $CD1a-$, $CD4+$ or $CD8+$).

How is the FLT3-ITD allelic ratio calculated and what is the threshold for 'High' risk?

The ratio is calculated as (Area under mutant peak / Area under wild-type peak). An allelic ratio $\ge 0.5$ is classified as High.

What is the molecular pathogenesis caused by the NPM1 exon 12 mutation?

The mutation replaces a nucleolar localization signal with a leucine-rich nuclear export signal, causing the persistent aberrant export of the NPM1 protein into the cytoplasm.

Which cytogenetic abnormality defines WHO Burkitt Leukemia/Lymphoma and what oncogene is involved?

It is defined by translocations involving the $MYC$ oncogene on chromosome $8$, most commonly $t(8;14)$.

What specific morphological feature characterizes AML with inv(16)?

The presence of dysplastic marrow eosinophils containing abnormal, giant purple-basophilic granules.