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Hemocytometer RBC and WBC Thoma pipet Unopette. Dilution problem Microscopy. Week 4: Hemocytometry. Red Cell and White Cell Counts. RBC problem: anemia, polycythemia WBC problem: leukocytosis, leukopenia RBC reference ranges Male4.50 - 6.00 x 10 6 /  L (x 10 12 /L)

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Week 4: Hemocytometry

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Week 4 hemocytometry l.jpg

Hemocytometer

RBC and WBC

Thoma pipet

Unopette

Dilution problem

Microscopy

Week 4: Hemocytometry


Red cell and white cell counts l.jpg

Red Cell and White Cell Counts

  • RBC problem: anemia, polycythemia

  • WBC problem: leukocytosis, leukopenia

  • RBC reference ranges

    • Male4.50 - 6.00 x 106/L (x 1012/L)

    • Female3.70 - 5.20 x 106/L (x 1012/L)

  • WBC reference range

    • Both5.0 - 10.0 x 103/L (x 109/L)


Diluting fluid l.jpg

Diluting Fluid

  • For RBC

    • Physiologic saline (0.8% NaCl)

    • Hayem’s solution

    • Reese-Ecker

  • For WBC

    • 3% Acetic acid

    • 0.1 N HCl


Thoma pipet l.jpg

RBC

Aspirate to 0.5 or 1.0

Dilute to 101

DF = 200 or 100

WBC

Aspirate to 0.5 or 1.0

Dilute to 11

DF = 20 or 10

Discard volume of one (1) in the stem

Thoma Pipet


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Improved Neubauer Hemocytometer


Hemocytometer chamber l.jpg

Hemocytometer Chamber

  • Depth = 0.1 mm

  • Grids

    • RBC use 5 small squares in the center large square

    • WBC use 4 corner large squares

    • Unopette for WBC use all 9 large squares


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Hemocytometer

Chamber


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Counting Rule

  • Do not count cells touching

    • Bottom line

    • Right line


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RBC Example…

  • Aspirate blood to 0.5, then dilute to 101

  • DF = 200

  • Counted 500 erythrocytes in 5 small squares

    RBC = 500 rbc x 25 Sm Sq x DF (200)

    5 Sm Sq 0.1 mm3

    = 5.00 x 106/mm3 (L)


Wbc example l.jpg

WBC Example…

  • Aspirate blood to 0.5, then dilute to 11

  • DF = 20

  • Counted 160 leukocytes in 4 large squares

    WBC = 160 wbc x 1 Lg Sq x DF (20)

    4 Lg Sq 0.1 mm3

    = 8.0 x 103/mm3 (L)


Unopette l.jpg

Unopette

  • Self-contained reservoir and 20 mL pipet

  • Ammonium oxalate diluent

  • DF = 100

  • Count leukocytes in all 9 large squares

  • Special formula to facilitate calculation


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Unopette


Unopette wbc example l.jpg

Unopette WBC Example…

  • Counted 70 leukocytes in 9 large squares

  • Add 10% of count

  • Multiply by DF

    WBC = (70 wbc + 7 wbc) x DF (100)

    mm3

    = 7.7 x 103/mm3 = 7.7 x 109/L


Compound microscope l.jpg

Compound Microscope

  • Ocular (adjustable binocular)

  • Mechanical stage

  • Condenser

  • Objectives (parfocal)


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Care of Microscopes

  • Carry with care

  • Protect long objective lens

  • Clean and disinfect stage

  • Wipe oil off OIL

  • Save lamp life


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