Differential Count and Blood Smear Preparation

Differential Count and Blood Smear Preparation

• A differential count, also known as a leukocyte differential count, peripheral differential, white blood cell morphology, or WBC differential, is formally known as a White Blood Cell Count Differential.
• It's typically part of a complete blood count (CBC).
• It determines the relative number of each type of white blood cell (WBC) in peripheral blood.
• Results are expressed as percentages (relative numbers of each WBC type in relation to the total WBC) and absolute values (percentage × total WBC).

Uses of Differential Count

• Assesses the body's ability to respond to and eliminate infections.
• Detects the severity of allergic and drug reactions.
• Detects the response to parasitic and other types of infections.
• Crucial for evaluating the reaction to viral infections.
• Evaluates response to chemotherapy.
• Identifies various stages of leukemia.

Manual Differential Count

• Approximates WBC count based on cell count/high-power field (hpf).
• WBC counts (per cubic millimeter) are estimated roughly as follows:
  • 2-4 cells/hpf = 4,000 – 7,000 WBC/mm³
  • 4-6 cells/hpf = 7,000 – 10,000 WBC/mm³
  • 7-10 cells/hpf = 10,000 – 13,000 WBC/mm³
  • 10-20 cells/hpf = 13,000 – 18,000 WBC/mm³

White Blood Cell Types

• Five basic WBC types: Neutrophils, Lymphocytes, Monocytes, Eosinophils, and Basophils.

Neutrophils

• Also known as Segmented Polymorphonuclear Leukocytes (PMNs), Neutrophils, or Segs.
• The most common WBC type, functioning as the primary defense against infection.
• Neutrophilia: Increased neutrophil count, typically a response to infection or severe injury (e.g., bacterial infection, appendicitis, chronic myelogenous leukemia).
• "Shift to the left": Increased band/stab cells in a blood smear, an early sign of a WBC response to an infection, even before the overall WBC count increases.

Eosinophils

• Play a role in allergic disorders and combating parasitic infections.
• Eosinophilia: Increased eosinophil count, often in response to allergic disorders, skin inflammation (e.g., scarlet fever), or parasitic infections; also seen in some bone marrow disorders such as eosinophilic leukemia.
• Eosinopenia: Decreased eosinophil count, often due to stress or steroid exposure.

Lymphocytes

• Play a role in both immediate and delayed responses to infections or inflammation.
• Lymphocytosis: Increased lymphocyte count, commonly seen in viral infections, infectious mononucleosis, lymphocytic leukemia, bone marrow cancers, or radiation therapy.

Monocytes

• Respond to inflammation, infection, and foreign bodies by ingesting and digesting foreign material.
• Monocytosis: Increased monocyte count, often seen in various kinds of infection and inflammatory disorders, including brucellosis and tuberculosis. Also associated with various cancers, including monocytic leukemia.
  • Other conditions associated with monocytosis: subacute bacterial endocarditis, typhoid fever, rickettsial infections, collagen diseases, Hodgkin’s disease, Gaucher's disease.

Basophils

• Basophilia: Increased basophil count, often seen in chronic inflammation, hypersensitivity reactions (e.g., food allergies), or leukemia.

Blood Smear Preparation

• Steps: Prepare blood smear, stain the smear, count the cells, and report results.
• Smear Types: Cover glass smear, wedge smear, spun smear, buffy coat smear, thick blood smear.
• Smear Preparation Methods: 2-Coverslip method (Ehrlich's method), Two-glass slide method (Wedge method), Glass slide - Coverslip method (Beacom's method), Spinner's method.
• Prerequisites: Clean slides & coverslips, appropriate blood drop size, quick smearing to prevent clotting, proper spreader angle & pressure.
• Factors Affecting Smear Thickness: Drop size, angle between slide & spreader, spreader speed, spreader pressure.
• Well-Stained Smear Features: Film length (2/3 to 3¼ slide length), finger-shape (feather edge), smooth edges, & "rainbow" appearance at thin edge when held to light. Whole blood drop picked up & spread.
• Coverslip vs Slide: "Perfect" coverslip smears are better for even distribution of leukocytes; however "perfect" slide smears easier to learn & average quality slide smears are better than those of average coverslips.
• Methods in Drying Blood Smear: Air drying, low flame (e.g., bunsen burner), using oven, methyl alcohol or absolute alcohol immersion (1-2 minutes).