Blood 2
Blood 2
Blood 2
APPARATUS:
1. Sterile lancet
2. Methylated spirit
3. Cotton wool
4. Automated micropipette
5. The Improved Neubauer Counting Chamber (Haemocytometer)
6. Cover slip
7. Diluting fluid ( 1% glacial acetic acid in distilled water + Gentian violet)
8. Microscope
Sterile Lancet
Methylated Spirit
Cotton wool
• AUTOMATED MICROPIPETTE
IMPROVED NEUBAUER COUNTING CHAMBER
((HAEMOCYTOMETER)
Cover Slips
DILUTING FLUID (Turk’s Solution)
MICROSCOPE
PROCEDURE
Ø Measure 0.38ml of diluting fluid and dispense into a small test
tube.
Ø Add 20µl (0.02ml) of well mixed EDTA anticoagulated venous
blood or free flowing capillary blood and mix.
Ø Assemble the counting chamber:
Make sure that the central grid area of the chamber and the
special haemocytometer cover glass are completely clean and
dry.
Slide the cover slip into position over the grid areas and press
down on each side until rainbow colours are seen.
ØCharge the hemocytometer with well mixed diluted
blood.
Ø Allow it to settle for 2 minutes.
Ø Using x 10 objectives lens, locate the 4 squares
(1,2,3,4).
Ø Count the total number of white cells in the 4 large
corner squares(4mm²).
Calculation of total WBC/cu.mm
= NO of cell counted × 1/Area counted × 1/depth
× D.F.
=No of cells counted × ¼ × 1/0.1 × 20
=No of cells counted × 50
Normal value:
Adult : 4.0 - 11.0 × 10⁹/L cells/L (4000 – 11,000
cells/mm³).
Neonates = 10.0 – 25.0 × 10⁹ cells/L ( 10,000 ̶
25,000cells/mm³).
CLINICAL SIGNIFICANCE
1. Microscope
3. Leishman’s stain
4. Electronic blood counter
MICROSCOPE
Microscope Glass Slide
LEISHMAN STAIN
ELECTRONIC BLOOD COUNTER
PROCEDURE
• Using a clean dry slides, drop a drop of blood on
the slide.
• Draw the spreader to touch the drop of blood.
Allow to extend along the edge of the spreader.
• Holding the spreader at the angle of 45°C, spread
the blood to make a film (⅔ of the slide). Not too
thick a film but smooth and not interrupted with
holes and ridges. It should have a tail at the end.
• Dry the slide.
• Stain with Leishman’s stain.
Preparation of slide.
STAINING PROCEDURE
5.MONOCYTES
• Largest white cell
• Nucleus often indented/ kidney shaped
• Cytoplasm is abundant, and has blue-gray appearance
• Vacuoles may be seen
• Phagocytic in function and are capable of injecting a number of bacteria.
• Normal range: 2 - 8 %.