Estimation of Hb in Human Blood

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The estimation of hemoglobin (Hb) in human blood is a crucial diagnostic test to assess the oxygen-carrying capacity of blood and identify conditions like anemia or polycythemia. Below is an overview of the methods, normal ranges, and clinical significance of hemoglobin estimation:

Normal Hemoglobin Levels

  • Men: 13.8–17.2 g/dL
  • Women: 12.1–15.1 g/dL
  • Pregnant Women: 11.0–14.0 g/dL
  • Children: 11.1–16.1 g/dL
  • Newborns: 14.0–24.0 g/dL

Methods of Hemoglobin Estimation

  1. Sahli’s Method (Acid Hematin Method)
    • Principle: Hemoglobin reacts with hydrochloric acid (HCl) to form acid hematin, which is visually compared against a standard color scale.
    • Procedure:
      1. Add 0.1N HCl to a graduated hemoglobin tube up to the mark.
      2. Add blood using a pipette.
      3. Mix and let it stand for 10 minutes.
      4. Compare the color with a standard comparator and note the hemoglobin level.
    • Advantages: Simple and inexpensive.
    • Disadvantages: Less accurate compared to modern methods.
  2. Cyanmethemoglobin Method (Drabkin’s Method)
    • Principle: Hemoglobin is converted to cyanmethemoglobin by reacting with Drabkin’s solution (containing potassium cyanide and ferricyanide). The absorbance is measured spectrophotometrically at 540 nm.
    • Procedure:
      1. Mix blood with Drabkin’s solution.
      2. Allow the reaction to stabilize.
      3. Measure absorbance using a spectrophotometer.
    • Advantages: Accurate and standardized.
    • Disadvantages: Requires specialized equipment.
  3. Automated Hematology Analyzers
    • Principle: Measures hemoglobin concentration using advanced optical or chemical methods.
    • Advantages: Fast, precise, and provides additional parameters like RBC count and hematocrit.
  4. Copper Sulfate Method
    • Principle: Blood is dropped into a solution of copper sulfate of known specific gravity. The blood sinks if hemoglobin concentration is adequate; otherwise, it floats.
    • Advantages: Quick screening, often used for blood donation.
    • Disadvantages: Qualitative, not quantitative.
  5. Tallqvist Method
    • Principle: Blood color is matched against a color scale on filter paper.
    • Advantages: Simple and portable.
    • Disadvantages: Least accurate.

Clinical Significance

  • Low Hemoglobin (Anemia):
    • Causes: Nutritional deficiencies (iron, B12, folate), chronic diseases, blood loss, bone marrow disorders.
    • Symptoms: Fatigue, pallor, shortness of breath.
  • High Hemoglobin (Polycythemia):
    • Causes: Dehydration, smoking, living at high altitudes, bone marrow disorders.
    • Symptoms: Headaches, dizziness, hypertension.
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