NEUBAUER CHAMBER (Haemacytometer)

1. NEUBAUER CHAMBER (Haemacytometer) Dr Maliha Sumbul

2. A device used for cell counting • Made of special optical glass • Used to count cells in suspension under a microscope • Improved Neubauer chamber most commonly used in haematology laboratory

3. BASIC DESIGN • Improved NC consists of a thick glass slide • Has 4 longitudinal grooves (depressions) in the central third of the slide – unfinished (white and granular) • Central support – smooth and polished • The ruled grid ( for counting ) is engraved on the central support – used as a counting chamber

4. Leukocyte Grid (FOR WBC) • Erythrocyte Grid (FOR RBC) • Thrombocyte Grid (FOR PLATELETS)

8. Haemacytometer

9. DEPTH METHOD OF CHARGING HOME WORK ?

10. 0.0025 mm2 = 0.2 divided by 4 = 0.05 x 0.05 = 0.0025 mm2 Area of ultra small square (square within RBC square)

12. Calculation factor Calculations of FACTOR • For WBC -------------- 50 • For RBC -------------- 10,000 • For PLATELET --------1,000

13. FORMULA FOR CELL COUNT • Cell count = N x Dilution Volume • Surface area = L X B • Volume = L X B X Depth

14. Dilution factor Dilutions: 380 ul 20 ul Total: 400 ul • WBC / PLT: 0.38 ml diluting fluid (3 %Glacial acetic acid) 0.02 ml Blood ( 1% AMMONIUM OXALATE) 0.40 ml TOTAL FOR DILUTION FACTOR: 0.40 = 20 0.02

15. 3980 ul 20 ul 4000 ul: total • FOR RBC: 3.98 ml Diluting fluid ( 0.9 % normal saline ) 0.02 ml blood 4.00 ml TOTAL FOR DILUTION FACTOR : 4.00 = 200 0.02

16. AFTER CARE OF THE CHAMBER

17. Possible sources of error • Inadequate mixing of specimen • Incorrect dilution • Chamber not clean • Cover slip not placed properly onto chamber • Air bubbles, fingerprints, oily surface or debris in chamber • Chamber overfilled • Not enough time for sedimentation of cells • Failing to count all the cells in the squares • Including artefacts in the count

18. QUESTIONS ?? Home Work Write a short note on optics and different parts of a MICROSCOPE

19. EXERCISES:QUESTION 1: • Calculate the Total WBC count provided: N = 102 ( Peripheral smear shows normal leukocyte count ) NO SHORT CUTS IN CALCULATIONS PLEASE

20. QUESTION 2: • Calculate the dilution factor for normal RBC count using Haemacytometer

21. QUESTION 3: • Calculate the Total Platelet count using the following information: N = 150 (The peripheral smear shows normal platelet count)

22. Assignment: • 0.38 ml 0.02 ml 1:20 0.40 ml • 0.78 ml 0.02 ml 1:40 0.80 ml • 0.18 ml 0.02 ml 1:10 0.20 ml