Erythrocyte & Leukocyte Disorders - Clinical Laboratory Science Chapter 10 Part THREE & FOUR

Erythrocyte & Leukocyte Disorders - Clinical Laboratory Science Chapter 10 Part THREE & FOUR

Microscopic Examination of Peripheral Blood

  • Preparation and Examination

    • Involves preparing, staining, and examining a thin smear of blood on a slide.

    • This examination is crucial for studying the morphology of red blood cells (RBCs), white blood cells (WBCs), and platelets.

  • WBC Differential

    • Determines the percentage of each leukocyte type through direct microscopic examination.

    • Manual review of the total blood count can confirm analyzer results, aiding in the diagnosis of blood disorders.

Microscopic Examination of Blood Film

  • Staining Technique

    • Morphology is studied by performing a Wright's-stained smear.

    • Examination should occur in the feather end (thin end) of the smear using a low power (10x) objective.

    • RBCs should be arranged such that they are just touching and not overlapping.

    • The film should ideally show many RBCs surrounding a lesser number of WBCs and platelets.

    • After selecting the appropriate area of the slide, switching to a 100X oil immersion lens allows for detailed examination of RBCs and platelets morphology.

Low-power (10X) & High-dry (40X) Objectives

  • Assessment of Blood Film

    • Evaluate the overall quality of the blood film and staining.

    • Estimate the concentrations of RBCs, WBCs, and platelets.

    • Identify the optimal area for WBC counting.

    • Scan for abnormal cells and clusters of platelets.

Oil-Immersion (100X) Objective

  • Technique for Examination

    • A drop of immersion oil must be placed on the selected area of the blood film, with direct contact between the oil and the lens necessary for optimal light focus onto the smear.

    • Examine RBC morphology for variations and abnormalities.

    • Estimate platelet count and morphology, as well as conduct a differential count of WBCs, examining for abnormalities in WBCs.

Normal Characteristics of Blood Cells

  • Normal RBCs

    • Most numerous type of blood cell, typically round in shape.

    • Anuclear: RBCs lose their nucleus before entering peripheral blood, leading to a biconcave disk shape.

    • Contains a rim of hemoglobin and an area of central pallor, measuring 6–8 µm.

    • Stain a reddish-orange color.

  • Normal Platelets

    • Smallest blood cell fragments; approximately 2-3 µm in diameter.

    • Formed from cytoplasmic fragments of megakaryocytes in bone marrow.

    • Cytoplasm stains blue and contains reddish-purple granules.

    • Typically round or oval with potential spiny projections, should not appear in clumps.

RBC Morphologic Examination

  • Examination focuses on the following variations:

    • Color or Staining Reaction Variations

    • Central pallor of the RBC should be approximately 1/3 of its diameter.

    • Pale RBCs show increased central pallor, termed hypochromic, indicating decreased hemoglobin content.

    • Spherocytes lack a center depression, appearing evenly stained throughout without central pallor.

    • Polychromasia refers to variation in staining color (pinkish blue), common in reticulocytes reflecting ribosomal RNA and hemoglobin levels in young RBCs.

    • Size Variations

    • Anisocytosis indicates increased variation in RBC size, quantifiable through RDW.

    • Microcytes (<78 fL MCV), associated with hypochromasia; present in conditions like iron deficiency, thalassemia, and lead poisoning.

    • Macrocytes (>100 fL MCV), observed in megaloblastic anemias due to vitamin B12 or folate deficiency.

    • Shape Variations (Poikilocytosis)

    • Varied RBC shapes found in various anemias and hemolytic states, with specific shapes indicating certain diseases.

Descriptions of Abnormal Cell Shapes

  • Acanthocyte: Spikes on cell membrane (spur cell)

  • Blister Cell: RBC with a bubble-like area on the membrane

  • Burr Cell (Echinocyte): Spiny projections distributed evenly

  • Crenated RBC: Scalloped edges, often due to incorrect drying during smear preparation

  • Elliptocyte/Ovalocyte: Oval-shaped RBC

  • Helmet Cell: Triangular shapes resembling a helmet

  • Schistocyte: Fragmented cells from mechanical trauma

  • Sickle Cell: Crescent-shaped with pointed ends

  • Spherocyte: Smaller, darker RBCs lacking biconcavity

  • Stomatocyte: Bowl-shaped with a mouth-like area in central pallor

  • Target Cell: RBCs exhibiting central and peripheral hemoglobin distribution, indicating a membrane defect

Red Cell Inclusions

  • Basophilic Stippling: Fine dark blue granules from ribosome and RNA precipitation in abnormal RBC formation.

  • Siderocyte (Pappenheimer Body): Blue-purple granules reflecting free iron presence, typically 1-2 per cell.

  • Howell-Jolly Bodies: Single purple granule derived from incomplete nucleus expulsion, notably present post-splenectomy.

  • Cabot Rings: Thread-like red-violet strands in reticulocytes, abnormal formations.

  • Nucleated RBCs: Normal in bone marrow, observed in peripheral blood during severe anemia.

Abnormal Red Cell Distribution

  • Rouleaux Formation: RBCs aggregate resembling stacked coins; clinically relevant in the thin smear area indicating increased fibrinogen.

  • Agglutination: Clumping caused by cold agglutinins, indicative of autoimmune hemolytic conditions or anemia.

RED BLOOD CELL MORPHOLOGY Chart

  • Overview of variations including size, shape, inclusions, and distributions with conclusional assessments (normal, mild, moderate, severe).

Platelet Estimation

  • Normal platelet estimation involves 6-20 platelets seen per 100x field; should be included with morphology for automation count verification.

  • Decreased counts can stem from conditions like radiation, drugs, chronic alcohol use, splenomegaly, or leukemia.

  • Increased counts may result from inflammatory responses, splenectomy, or refer from other blood disorders.

Characteristics of White Blood Cells (WBCs)

  • WBCs are the largest blood cells, ranging from 5-20 µm depending on type.

  • Types of WBCs:

    • Granulocytes: Comprising neutrophils, eosinophils, and basophils, identifiable by segmented nuclei and granules.

    • Agranulocytes: Consist of lymphocytes and monocytes characterized by large staining nucleuses with varying cytoplasm.

    • Characteristics for differentiation include size, nuclear shape and characteristics, and cytoplasmic features (such as color and granule presence).

WBC Differential Techniques

  • Involves identifying and counting a minimum of 100 WBCs (50 if WBC count <1 x 10^9/L).

  • Report WBC as absolute values based on counts made.

Immature vs. Mature WBCs

  • Size: Immature WBCs are larger than their mature counterparts.

  • Nuclear Characteristics: Immature cells possess larger and rounder nuclei; mature cells have lobular/nuclear indentations.

  • Staining Patterns: Immature cells display reddish-violet while mature cells are basophilic.

  • Functions: Immature cells denote mitotic figures while mature WBCs have functional cytoplasmic inclusions for phagocytosis.

Quantitative Changes in WBC Counts

  • Leukocytosis: An increase in WBC count.

  • Leukopenia: A decrease in WBC count.

  • Specific changes in cell types can indicate conditions:

    • Neutrophilia: Increase typically in acute infections, chemical exposures, etc.

    • Neutropenia: Increased infection risk, commonly due to chemotherapy or inherited disorders.

    • Eosinophilia: Commonly associated with allergic reactions, while eosinopenia indicates hyperadrenalism.

    • Basophilia: Associated with chronic myelogenous leukemia and other conditions.

Toxic Changes in WBCs

  • Dohle Bodies: Clear, blue-staining areas in neutrophil cytoplasm indicating immature stages related to infections or burns.

  • Toxic Granulation: Enhanced, larger granules in neutrophils due to acute infections or drug poisoning.

  • Toxic Vacuolation: Presence of vacuoles in the cytoplasm indicating severe acute bacterial infections.

Clinical Considerations

  • Both acute and chronic leukemias can demonstrate abnormal WBC counts.

  • Acute Leukemia: Characterized by an influx of immature blast cells alongside elevated WBC counts.

  • Chronic Leukemia: Presents mature WBCs with counts variable from normal to extremely high (>50,000 cells).

Conclusion

  • Comprehensive understanding of RBC and WBC morphology, distributions, and disorders enhances diagnostic accuracy in clinical laboratory settings.

  • Detailed examination methods and knowledge of variations in blood cells are vital for proper interpretation and treatment.