Study Myeloperoxidase Staining Enhances Blood Disease Diagnosis

Myeloperoxidase (MPO) staining, also known as leukocyte peroxidase (POX) staining, is a widely used cytochemical staining method. Primarily employed for staining bone marrow cells and blood smears, this technique detects intracellular peroxidase activity to assist in diagnosing hematologic disorders, particularly in leukemia classification.

MPO staining relies on the catalytic reaction of intracellular peroxidase. This enzyme breaks down oxides to produce oxygen, which then reacts with potassium iodide to form iodine. The iodine combines with Wright-Giemsa dye, creating colored granules in the cytoplasm that reveal peroxidase distribution under microscopy. Different cell types exhibit varying peroxidase content and distribution patterns, allowing MPO staining to distinguish between them.

Standard MPO staining requires specialized reagent kits to ensure accuracy and reliability. A typical kit contains:

• Solution A: Eosin solution - An acidic dye that stains cytoplasm red or pink
• Solution B: Azure II - A basic dye that stains nuclei blue or purple
• Solution C: Potassium iodide PBS buffer - Contains the peroxidase reaction substrate and maintains pH stability
• Solution D: Wright-Giemsa stain - A dual-purpose stain that enhances cellular morphology

Kits are available in various sizes (5-test, 20-test, and 100-test configurations) to accommodate different laboratory needs.

1. Working solution preparation: Mix Solution C with Solution D (typically 1.0ml:250µl ratio). Use within two hours.
2. Smear fixation: Fix prepared bone marrow or blood smears with methanol or ethanol for 5-10 minutes.
3. Staining: Immerse fixed smears in working solution for 5-10 minutes, controlling temperature and humidity.
4. Rinsing: Gently wash with distilled water or PBS buffer to remove excess stain.
5. Counterstaining (optional): Use hematoxylin to enhance nuclear contrast.
6. Drying: Air-dry or use a blow dryer.
7. Microscopy: Examine stained smears to evaluate peroxidase activity and distribution.

Result interpretation requires specialized expertise. Common findings include:

• Positive reaction: Red-brown to dark blue cytoplasmic granules (weak vs. strong positivity). Granules may cover the entire cytoplasm or obscure the nucleus.
• Negative reaction: Blue cytoplasm without granules and uniformly purple-red nuclei.
• Eosinophils: Exhibit intense dark blue staining, sometimes with extracellular needle-like crystals.

• Leukemia classification: Distinguishes myeloid (MPO-positive) from lymphoid (MPO-negative) leukemia, guiding treatment decisions.
• Acute myeloid leukemia (AML) diagnosis: Confirms myeloid origin and aids subtyping, with varying positivity rates indicating different AML subtypes.
• Myelodysplastic syndrome (MDS) evaluation: Assesses cellular maturation and differentiation, with abnormal MPO patterns suggesting MDS.
• Other hematologic disorders: Useful in diagnosing chronic myeloid leukemia and myelofibrosis.

• Reagent quality and storage conditions
• Strict protocol adherence
• Proper smear preparation
• Optimal staining duration
• Expert microscopic evaluation by pathologists

Technological advances continue to enhance MPO staining. Immunohistochemistry now combines MPO with immune markers for greater precision, while flow cytometry enables quantitative MPO detection. As this classic cytochemical method evolves, it promises to deliver even greater contributions to hematologic diagnosis and treatment.