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Hematology

Hematocrit

Overview of Hematocrit (HCT)

  • Definition: The hematocrit is the percentage of blood volume occupied by red blood cells (RBCs). It’s a simple but valuable measurement in assessing a patient’s red blood cell status
  • Alternate Names: Packed Cell Volume (PCV)
  • Clinical Significance
    • Detection and Monitoring of Anemia: Low HCT indicates anemia (decreased red blood cell mass)
    • Detection of Polycythemia/Erythrocytosis: High HCT indicates increased red blood cell mass
    • Assessment of Dehydration: Can be elevated in dehydration due to decreased plasma volume
    • Monitoring Fluid Balance: Helps assess changes in blood volume
  • Methods
    • Manual Methods: Centrifugation of blood in a capillary tube (microhematocrit method)
    • Automated Methods: Calculated by automated hematology analyzers based on RBC count and MCV

Manual Microhematocrit Method

  • Principle

    • Whole blood is collected in a capillary tube coated with an anticoagulant (usually heparin)
    • The tube is centrifuged to pack the red blood cells at the bottom of the tube
    • The height of the packed red blood cell column is measured and compared to the total height of the blood column to determine the hematocrit percentage

  • Materials

    • Capillary Tubes:
      • Glass or plastic capillary tubes (typically 75 mm long with an internal diameter of 1-2 mm)
      • Coated with an anticoagulant (heparin)
    • Centrifuge:
      • A specialized centrifuge designed to hold and spin capillary tubes at high speeds
      • Should be capable of generating a relative centrifugal force (RCF) of at least 10,000 g
    • Reader:
      • A device for accurately measuring the height of the packed red blood cell column and the total blood column in the capillary tube
      • Can be a microhematocrit reader card or a digital reader
    • Sealing Clay:
      • Used to seal one end of the capillary tube after filling

  • Procedure

    1. Sample Collection:
      • Collect whole blood in an EDTA (purple-top) tube
      • Capillary blood (fingerstick or heelstick) can also be used, but is less accurate
    2. Filling the Capillary Tube:
      • Mix the blood sample thoroughly
      • Hold the capillary tube horizontally and touch one end to the blood sample
      • Allow the tube to fill by capillary action to at least 2/3 of its length
      • Avoid introducing air bubbles into the tube
    3. Sealing the Tube:
      • Wipe the outside of the capillary tube clean
      • Seal one end of the tube with sealing clay to prevent leakage during centrifugation
    4. Centrifugation:
      • Place the filled and sealed capillary tubes into the microhematocrit centrifuge
      • Centrifuge for the recommended time (typically 5 minutes) at the specified RCF (10,000-12,000 g)
    5. Reading the Hematocrit:
      • Remove the capillary tubes from the centrifuge
      • Carefully place the tube on the microhematocrit reader
      • Align the bottom of the packed red blood cell column with the 0% line on the reader
      • Align the top of the plasma layer with the 100% line on the reader
      • Read the hematocrit value at the interface between the packed red blood cells and the plasma
    6. Record and Report Results:
      • Record the hematocrit value as a percentage (%)
      • Report the result along with the reference range
      • Perform duplicate measurements and average the results (if duplicate results are within acceptable limits)

  • Sources of Error in Manual Hematocrit Measurement

    • Improper Sealing of the Capillary Tube: Can lead to leakage and falsely low results
    • Inadequate Centrifugation: Insufficient packing of red blood cells can lead to falsely low results
    • Delay in Reading: Prolonged delay after centrifugation can lead to cell settling and inaccurate readings
    • Air Bubbles: Air bubbles in the capillary tube can interfere with the reading
    • Incorrect Use of the Reader: Not aligning the tube properly on the reader can lead to inaccurate results
    • Excess Anticoagulant: Overfilling a capillary tube with a blood sample in an anticoagulant collection tube is not recommended

Automated Hematocrit Measurement

  • Principle

    • Automated hematology analyzers typically calculate the hematocrit based on the red blood cell count and the mean corpuscular volume (MCV)
    • Formula: HCT = (RBC Count x MCV) / 10

  • Procedure

    • The automated analyzer aspirates a small amount of the whole blood sample
    • The RBC count and MCV are measured using electrical impedance, optical scatter, or other methods
    • The hematocrit is calculated using the formula above

  • Advantages

    • Increased Speed and Efficiency
    • Improved Accuracy and Precision
    • Reduced Labor Costs

  • Limitations

    • Can be affected by certain interfering substances or conditions (see below)
    • Does not directly measure the packed cell volume; it is a calculated value

Interfering Substances & Conditions

Several factors can interfere with both manual and automated hematocrit measurements, leading to inaccurate results. It’s crucial to be aware of these interferences and take appropriate corrective actions:

  • Lipemia
    • Turbidity from high lipid content can falsely elevate the hemoglobin reading, which can affect the calculated hematocrit
    • Corrective Action: Perform a saline replacement procedure or use a lipemia clearing agent
  • High White Blood Cell Count (Leukocytosis)
    • Extreme leukocytosis can cause falsely elevated hemoglobin and hematocrit readings
    • Corrective Action: Dilute the sample and multiply the results by the dilution factor
  • Cold Agglutinins
    • These antibodies can cause RBCs to clump together, leading to inaccurate cell counts and hematocrit measurement
    • Corrective Action: Warm the sample to 37°C to dissociate the agglutinins
  • Abnormal Red Blood Cell Size or Shape
    • Spherocytes, schistocytes, and other abnormally shaped RBCs can affect the accuracy of both manual and automated hematocrit measurements
  • High Hemoglobin (Results beyond the linearity of the instrument)
    • Dilute the blood so that the Hgb result is within the linear range of the instrument

Quality Control (QC)

  • Calibrate the Instruments

    • Perform regular calibration of the microhematocrit centrifuge and automated hematology analyzer according to the manufacturer’s instructions

  • Run Control Materials

    • Run control materials at regular intervals (e.g., daily, with each batch of samples)
    • Use controls that are appropriate for the types of samples being analyzed (e.g., whole blood controls)

  • Review Control Results

    • Review control results and evaluate them using statistical methods (e.g., Levey-Jennings charts, Westgard rules)
    • Take corrective action if control results are outside the acceptable range

  • Check the Rule of Three

    • Verify that the hemoglobin, hematocrit, and RBC count are consistent with each other, using the “Rule of Three”:
      • HGB x 3 = HCT (± 3)
    • If the Rule of Three is not met, investigate potential errors

Reporting Results

  • Report the hematocrit value as a percentage (%)
  • Include the reference range for the patient’s age and sex
  • Note any abnormal results or flags
  • Document all quality control procedures and corrective actions taken