1. WBC manual count using hemocytometer

2. Introduction Blood represent about 8% of total body weight
It consist of 3 type of specialized cellular elements:
Erythrocytes RBCs
Leukocytes WBCs
Platelets (thrombocytes)
These cells suspended in complex liquid plasma

3. Introduction Blood perform two major function:
Transport through the body
O2 & CO2
Food molecules (Glucose, lipid, aa)
Wastes (ex. Urea)
Hormones
heat
Defense of the body against infections & other foreign materials, all WBCs participate in these defenses

4. Introduction
All various types of blood cells are produced in the bone marrow

5. Introduction WBCs (leukocytes) Have nuclei Consist of:
lymphocytes with relatively clear cytoplasm
And 3 types of granulocytes whose cytoplasm contain granules

6. Introduction
WBCs count is the count of leukocytes in a volume of blood
Expressed as WBCs/mm³

7. Principle
The # of WBCs is very large, so its practical to dilute a sample with diluting solution (2% Glacial acetic acid with methylene blue)
This solution will lyses cytoplasmic membrane, and leave the nuclei of WBCs.

8. Material Blood sample (EDTA anticoagulated blood or capillary blood)
WBCs diluting pipette
Diluting solution (2% AA with methylene blue)
Hemocytometer
microscope

9. The Hemacytometer contains 2 Neubauer counting chamber
Each chamber contains:
*4 WBC counting squares
*Each contains 16 squares

10. The Hemacytometer

11. The Hemacytometer
]0.25mm
]0.20mm

12. Methodology
With a safety bulb draw blood up to 0.5 marks on WBCs pipette and complete to 11 with WBCs diluting solution.
Mix for 2-3 minute.

13. Charge hemacytometer
Discard the first 4-5 drops
Place tip of the pipette at the edge central platform
Then let the hemacytometer to stand on the bench for 3-5 minute.

14. Count and calculate:

15. Count and calculate:

16. Calculation Number of cells/mm³= counted cells in 4 large square x
diluting factor x
volume correction factor.

17. Calculation The dilution factor= total volume/sample volume
= 11-1/0.5 =20
Volume correction factor=
Desired volume/counted volume
1 mm³/ counted volume

18. Calculation counted volume = The total volume of the 4 large squares=
= Volume x number of large squares
= (width x length x depth )x 4
= (1mm x 1mm x 1/10 mm) x 4 =0.4mm³
Volume correction factor=
1 mm³/ counted volume = 1 /0.4 = 2.5

19. Calculation Number of cells/mm³= counted cells in 4 large square x
diluting factor x
volume correction factor.
Number of cells/mm³= n x20 x 2.5
= n x 50

20. Example: If total # of WBCs in 4 squares is 120
Then the # of WBCs in 1mm³=
120 x 50 =6000

21. Normal values of WBCs: Newborn 9000-30000 cell/mm³
Adult cell/mm³

22. Significance of the test
Leukocytosis
Occur as an indicator of body defense against foreign materials (bacteria, parasites, toxins)
Metabolic disorders
Chemical and drug poisoning
Acute hemorrhage

23. Significance of the test
Leukopenia
Result from X-ray therapy
Alcoholism
Antibiotic therapy
Typhoid infection
Measls
Infectious hepatitis TB
And cirrhosis of the liver

24. Sources of error Flooding of chamber with excess sample
Failing to count all the cells in the squares or conversely including artifacts in the count.