Macro Method (Wintrobe Method) for Estimation of Packed Cell Volume (PCV) or Hematocrit (HCT)

Hematocrit (HCT), also known as packed cell volume (PCV) or erythrocyte volume fraction, measures the proportion of red blood cells in a sample of anticoagulated blood after centrifugation. This measurement indicates the relative amount of red blood cells compared to plasma. Typically expressed as a percentage of the total blood volume or as a decimal fraction, a hematocrit value of 40% means that 40 milliliters of red blood cells are present in 100 milliliters of blood. This metric is essential for diagnosing and monitoring various medical conditions. An elevated hematocrit can suggest an increased number of red blood cells or a reduction in plasma volume, such as in dehydration. Conversely, a decreased hematocrit may indicate anemia, characterized by reduced red blood cell production, increased red blood cell destruction, or dilution of blood cells due to an increased plasma volume, commonly seen during pregnancy or in conditions causing hemodilution.

Principle of Hematocrit (Packed Cell Volume) Measurement

Anticoagulated whole blood is centrifuged in a Wintrobe tube to fully pack the red blood cells. The volume of these packed cells is directly read from the tube. An advantage of this method is that it allows for the erythrocyte sedimentation rate test to be set up prior to performing the packed cell volume (PCV) measurement.

Essential Equipment for Measuring Packed Cell Volume (PCV)

1. Wintrobe Tube: This tube is approximately 110 mm in length, marked at 1 mm intervals up to 100 mm. It has an internal diameter of 3 mm and can hold around 3 ml of blood.
2. Pasteur Pipette: Equipped with a rubber bulb and a capillary long enough to reach the bottom of the Wintrobe tube.
3. Centrifuge: Capable of reaching a speed of 2300 g.

Specimen Requirements for Hematocrit (HCT) Test

Venous blood should be collected in EDTA (1.5 mg EDTA per 1 ml of blood) or in double oxalate. The test must be performed within 6 hours of blood collection.

Method for Determining Hematocrit (HCT) or Packed Cell Volume (PCV)

1. Mix the anticoagulated blood sample thoroughly.
2. Use a Pasteur pipette to draw the blood sample, introducing the pipette to the bottom of the Wintrobe tube. Fill the tube up to the 100 mark, ensuring the tip of the pipette remains under the rising meniscus to prevent foaming.
3. Centrifuge the sample at 2300 g for 30 minutes. To balance the centrifuge, place a second Wintrobe tube filled with blood from another patient or with water.
4. Read the length of the red cell column.

Hematocrit can be expressed either as a percentage or as a fraction of the total blood volume.

Clinical Significance of Packed Cell Volume (PCV) and Hematocrit (HCT)

PCV is below the normal range in cases of anemia. It increases in conditions such as dehydration, shock, burns, and polycythemia. After centrifugation, three distinct zones are visible in the Wintrobe tube from top to bottom: plasma, a buffy coat layer (a thin, grayish layer of white cells and platelets, about 1 mm thick), and packed red cells.

Normal plasma appears straw-colored. It is colorless in iron deficiency anemia, pink in hemolysis or hemoglobinemia, and yellow in the presence of elevated serum bilirubin (jaundice). In hypertriglyceridemia, plasma appears milky. An increased thickness of the buffy coat layer occurs when there is a rise in the number of white cells or platelets, as seen in leukocytosis, thrombocytosis, or leukemia. Smears from the buffy coat can be used to demonstrate lupus erythematosus (LE) cells, malaria parasites, or immature cells.