It is necessary to determine haematocrit with 6 hours after the blood collection.
Principle:
Blood is centrifuged in sealed capillary tube and PCV is determined by a special haematocrit reader.
Requirements: Haematocrit Centrifuge:
1. It runs at high speed and produces RCF of 12,000 X g and runs at a speed of about 15,000RPM. It contains a head to hold the capillary tubes.
2. Haematocrit Reader this is supplied by the manufacturer. It also can be made by hand.
3. Capillary Haematocrit tubes these are 75 mm in length with approximately 1 mm diameter.
4. Soft wax or modeling caly. This is used to seal the end of the haematocrit tube.
Procedure:
1. Draw the specimen into an appropriate capillary tube. Fill in the tube to about ¾ length.
2. Seal both the ends of the tube with soft wax or modeling clay. It is plugged to a depth of about 1 cm.
3. Write identification number on the tube by using a marking pencil.
4. Place the tube with another similar tube in the radial groves of the centrifuge head exactly opposite to each other (empty capillary tube also can be used.)
5. Close the centrifuge cover and centrifuge the tubes at high speed (about 15,000 RPM) for 5 minutes.
6. Remove the capillary tube. It will show three layers (a) clear plasma at the top (b) whitish buffy coat at the middle and (c) column or red cells at the bottom.
7. Hold the tube against the haematocrit scale until the line marked 1.0 passes through the top of the plasma column.
8. Move the tube across the scale until the line marked 1.0 passes through the top of the plasma column.
9. The line that passes through the top of the column of red cells gives the value of PCV (haematocrit).
Source of Error:
1. Haemolysed specimen will yield false low values.
2. Inadequate mixing of blood and incompleteness of packing may lead to erroneous results.
Disadvantage:
It requires a special centrifuge and disposable capillary tubes.
Additional Observations:
Note any abnormal findings such as:
1. The colour of plasma (yellow for jaundice and reddish for haemolysis).
2. Increased buffy coat for increased white blood cells.
Macro Method:
PCV – Packed Cell Volume
Principle:
When, anti-coagulated blood is centrifuged in a haematocrit tube at high speed, the erythrocytes sediment at the bottom. The red cell column is called as packed cell volume (PCV) or haematocrit (cell volume percent.)
Requirements:
1. Specimen:
EDTA or double oxalated blood (The specimen need not be fasting sample).
2. Wintrobe Haematocrit Tube:
It is 110 mm long tube with a 2 mm internal bore. It is graduated from 0 to 100 mm (10 cm), the scale with descending order is used for PCV determination.
3. Pasteur pipette or a syringe with needle.
4. Centrifuge
Procedure:
1. Mix the blood sample carefully
2. Label a wintrobe tube.
3. Fill the tube by using Pasteur pipette or a syringe up to the 100 mark (use a needle of wide bore to prevent mechanical breakdown of red blood cells). Avoid trapping of air bubbles.
4. Place the tube in a centrifuge cup (use another empty wintrobe tube to fill the opposite cup or a tube filled with another specimen to balance.
5. Centrifuge for 30 minutes at 3000 RPM.
6. Note the reading. Multiply by 100 for volume percent. If the blood is above the 10 mark calculate the cell pack by dividing the height of the column eight of the cell column and plasma and then multiply by 100.
7. Make the following Observations:
a. Color and Opacity of Plasma Expected Condition:
i. Yellow: May be jaundice
ii. Milky: Lipaemia
iii. Cloudy: May be multiple myeloma.
iv. Reddish: Haemolysis.
b. Buffy Layer:
Normally it is 0.5 to 1 mm. Each 0.1 mL = 1000 cells per cubic mn (µl)
Approximately
Precaution:
It is necessary to wash wintrobe tubes immediately after the test by introducing a thick steel wire repeatedly in the tube under running water.