Total leucocyte count

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  • 1. TOTAL LEUCOCYTE COUNT Mr Govardhan Joshi
  • 2. Principle • The sample of blood is diluted with a turk’s fluid which destroy RBC and stain the nuclei of leucocyte. The cell are then counted in a calibrated chamber and their number in undiluted blood reported as leucocyte/cumm.
  • 3. Requirement • • • • • • Counting chamber WBC count Turk’s fluid EDTA blood Microscope Timer
  • 4. Turk’s fluid Composition: • Glacial acetic acid= 1.5ml→lyses RBC • 1% gention violet= 1ml→ stain nuclei of WBC • Distilled water= upto 100ml→ solvent • Thymol= 1 pinch→prevent growth of fungus The diluting fluid is not isotonic and the very dilute acid solution lyses RBC but not WBC
  • 5. Methods • Microdilution method • Bulk dilution method
  • 6. Procedure microdilution method • Draw the well mixed anticoagulated blood upto mark 0.5 • Clean the tip of pipette and adjust the level exactly upto mark 0.5 • Draw diluting fluid upto mark 11 • Mix well by rotating pipette for 2-3 minutes • Charge the neubauer chamber by discarding 1-2 drops • Allow the cell to settle down for 2 minutes • Count WBC under low power(10x) in four large corner square
  • 7. Procedure for bulk dilution method • Mix 380µl of diluting fluid and 20µl of blood in a clean test tube • Other steps same as above
  • 8. Calculation WBC count=N x dil. Factor x depth factor/area counted WBC count= N x 50
  • 9. In case of leucopenia • Dilution→ 1:10 In case of leucocytosis • Dilution→ 1:100
  • 10. Normal values Adult= 4,000- 11,000/cumm Birth= 10,000- 25,000/cumm 1-3yrs= 6,000- 18,000/cumm 4-7yrs= 6,000- 15,000/cumm 8-12yrs= 4,500- 13,500/cumm
  • 11. Clinical significance • • • • Leukocytosis Leukaemia Myeloproliferative disorder Infection Physiological: age, muscular exercise etc • • • • • Leucopenia Typhoid Aplastic anemia Dengue Influenza Malaria
  • 12. Corrected total WBC count for nucleated RBC • Done when nucleated RBC are seen in peripheral blood smear • Corr. WBC count= 100x Uncorrected WBC/100+NRBC Uncorrected WBC= total nucleated cell counted NRBC= nucleated RBC seen in PBS/ 100 WBC
  • 13. or Corr. WBC= Uncorr.WBC-(NRBCx Uncorr.WBC) 100+NRBC