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Agile - affordable - accurate What happens in the aperture? Basics of Hematology cell counting.

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Presentation on theme: "Agile - affordable - accurate What happens in the aperture? Basics of Hematology cell counting."— Presentation transcript:

1 agile - affordable - accurate What happens in the aperture? Basics of Hematology cell counting

2 agile - affordable - accurate Blood cells NameRBCWBCPLT Population million/µl /µl /µl Shape, diameter Biconcave shape Diameter 8 µm Thickness 2 µm NE,EO,BA 15 µm LYM 8-15 µm MON µm Fragments Diameter 2-4 µm Mean volume 90 fl various 12 fl

3 agile - affordable - accurate Blood cells – under microscope NEU EO LYM BASO MONO NRBC PLT RBC

4 agile - affordable - accurate Hematology parameters 1 White blood cell – WBC (pcs/l, pcs/µl) Number of leukocytes. WBC = WBC cal x (pcs/l or pcs/µl) Red Blood Cell – RBC (pcs/l, pcs/µl) Number of erythrocytes. RBC = RBC cal x (pcs/l or pcs/µl) Haemoglobin concentration - HGB (g/dl, g/l, mmol/l) Measured photometrically at 540 nm; in each cycle blank measurement is performed on diluent HGB = HGB cal x (HGB measured – HGB blank ) Mean Corpuscular Volume - MCV (fl= l) Average volume of individual erythrocytes derived from the RBC histogram. Haematocrit – HCT (percentage, absolute) Calculated from the RBC and MCV values. HCT percentage = RBC x MCV x 100, HCT absolute = RBC x MCV Mean Corpuscular Haemoglobin – MCH (pg, fmol) Average hemoglobin content of erythrocytes, calculated from RBC and HGB values. MCH = HGB / RBC

5 agile - affordable - accurate Hematology parameters 2 Mean Corpuscular Haemoglobin Concentration – MCHC (g/dl, g/l, mmol/l) Calculated from the HGB and HCT values. MCHC = HGB / HCT absolute Unit of measurement is displayed according to the one chosen for HGB result (g/dl, g/l or mmol/l) Red cell Distribut. Width - RDW-SD and Platelet Distribut. Width - PDW-SD (fl) Red cell Distribut. Width - RDW-CV and Platelet Distribut. Width - PDW-CV (absolute) The distribution width of the erythrocyte or platelet population derived from the histogram at 20% of peak xDW-SD = RDW cal x (P2 – P1) (fl), xDW-CV = RDW cal x 0.56 x (P2 – P1) / (P2 + P1) by the factor of 0.56 CV is corrected to the 60% cut

6 agile - affordable - accurate Hematology parameters 3 Platelet – PLT (pcs/l, pcs/µl) Number of thrombocytes (platelets) PLT = PLT cal x (pcs/l or pcs/µl) Mean Platelet Volume – MPV (fl) Average volume of individual platelets derived from the PLT histogram. Thrombocrit – PCT (percentage, absolute) Calculated from the PLT and MPV values. PCT percentage = PLT x MPV x 100, PCT absolute = PLT x MPV White blood cell differential: LYM#, LY% : lymphocytes MID#, MID%: monocytes and some eosinophil granulocytes GRA#, GR%: neutrophil, eosinophil and basophil granulocytes Absolute values counted in the channels determined by the three WBC discriminators: Percentages calculated from the absolute WBC value. 1.: RBC-LYM discriminator 2.: LYM-MID discriminator 3.: MID-GRA discriminator

7 agile - affordable - accurate Blood sample

8 agile - affordable - accurate Impedance cell counting principle dR/R = 1 / = 50ppm, RBC = 1mV, min PLT= 40 µ V

9 agile - affordable - accurate Aperture clogging effect

10 agile - affordable - accurate Coincidence effect >1 cells in aperture look one big cell Less counts, distorted histogram Solution: diluting samples, mathematical coincidence correction Linearity range (80um): WBC: 100 x 10^3/ul RBC: 10 x 10^6/ul PLT: 1000 x 10^3/ul

11 agile - affordable - accurate Sample preparation: diluting, lysing 1. Blood 3. Hemolyser 1: : a b c 1.a + diluent b 2.b + diluent c RBC/PLT 3.b + Lyser WBC/HGB

12 agile - affordable - accurate Differential Lysing process PLT & RBC LYM MON GRAN RBCs destroyed WBCs selectively shrank to nuclei

13 agile - affordable - accurate 3-part Differential WBC Histogram & HGB

14 agile - affordable - accurate Hemoglobin measuring principle Specific wavelength light source – green 560 nm Light to frequency converter Ambient light compensation with U/D counting, and light chopping LED SAMPLEDETECTORLED switching: Frequency output

15 agile - affordable - accurate Hemoglobin calculation – HGB blank HGB needs blank measurement on clean diluent to compensate temperature drift HGB = C * ln (CNT blank / CNT sample ) CNT blank = HGB count on diluent = CNT sample = HGB count on sample = 4.000

16 agile - affordable - accurate Thank you for your attention!

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