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Electrical Impedance and Colorimetric Measurements. Joanna Ellis, MLS(ASCP). Objectives. Cite the electrical impedance principle of cell counting. Identify and interpret microcytic and macrocytic RBC histograms Define coincidence

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Electrical Impedance and Colorimetric Measurements

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Electrical impedance and colorimetric measurements l.jpg

Electrical ImpedanceandColorimetric Measurements

Joanna Ellis, MLS(ASCP)

Objectives l.jpg


  • Cite the electrical impedance principle of cell counting.

  • Identify and interpret microcytic and macrocytic RBC histograms

  • Define coincidence

  • Identify the cell populations represented on a 3-part differential WBC histogram

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HistoryYou CAN patent a hole

  • Prior to the 1950s blood cell counts were performed by manual methods:

    • Hemacytometer blood counts

    • Spun hematocrits

    • Spectrophotometrically determined hemoglobins

    • Peripheral blood cell evaluation for all differentials

  • In 1953, Wallace Coulter patented the Coulter Principle in which particles are counted in fluid passed through a hole. The incredulous attorneys who had told him “You can’t patent a hole” were proven wrong.

  • Hematology automation has since grown to include additional principles such as optical light scatter and flow cytometry.

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The Coulter Principle

  • The poorly conductive blood cells are suspended in a conductive diluent (liquid).

  • The diluent is passed through an electric field created between two electrodes.

  • The liquid passes through a small aperture (hole).

  • The passage of each particle through the aperture momentarily increases the impedance (resistance) of the electrical path between the electrodes.

  • The increase in impedance creates a pulse that can be measured.

  • The number of pulses = blood cell count

  • The amplitude (height) of the pulse = Volume of cell

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Electrical Impedance (The Coulter Principle)

Sweep Flow: Steady stream of diluent that flows behind each aperture to prevent cells from re-entering the aperture

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Counting Chambers

  • Most common chambers using impedance:

    • RBC/Platelet chamber

    • WBC chamber

RBC/Platelet Chamber

WBC Chamber

Differential Chamber

Reticulocyte Channel

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RBC/Platelet Chamber

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RBC and Platelet Histograms

The black line represents normal cell distribution. The red line on the RBC histogram graphically represents a microcytic red cell population.

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Bimodal Histogram

  • Bimodal peak can be seen in situations such

  • Cold agglutinin disease

  • Post-transfusion

  • Post-treatment of IDA

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Coincidence: Multiple cells passed through the aperture at once.

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WBC Chamber


Lysing agent

Aperture in electric field

Hgb released

In some analyzers the WBC count is directly measured by electric impedance after the red cells have been destroyed by a lysing agent. The lysing agent also shrinks the leukocyte cell membrane and cytoplasm; therefore, the WBC count represents the measure of the cell volume not native cell size. Colorimetric measurements are used to determine hemoglobin.

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Colorimetric Measurements

  • Hemoglobin is often determined by a colorimetric method.

    • Imidazole

      • Non-cyanide reagent with color change and read at 540nm

      • Instruments

        • Abbott CELL-DYN Sapphire

    • Sodium Lauryl Sulfate

      • Non-cyanide reagent with color change and read spectrophotometrically

      • Instruments

        • Sysmex XT and XE

    • Lysing agent converts free hemoglobin to cyanmethemoglobin and read spectrophotometrically at 540nm.

      • Instruments

        • Advia 120

        • Some Beckman Coulters

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Three-part Differential WBC Histogram

Lymphocytes: 35-92L

Mononuclear Cells: 92-152 fL

Granulocytes: 152-450 fL

References l.jpg


  • "Abaxis Veterinary Diagnostics - VetScan HM2 Technology." Abaxis Medical Diagnostics - Redirect. Abaxis Medical Diagnostics, 2006. Web. 12 Sept. 2010. <http://www.abaxis.com/veterinary/vetscan_hm2_technology.html>.

  • Graham, Marshall Don. "The Coulter Principle: Foundation of an Industry." The Association for Laboratory Automation :: Home. JALA Volume 8, Issue 6, Dec. 2003. Web. 12 Sept. 2010. <http://labautomation.org>.

  • Kelly, Kathleen. "Modules :: CLIA :: CME." Hematology in the Physician Office Laboratory Section I. University of Iowa Carver College of Medicine, 2008. Web. 12 Sept. 2010. <http://www.medicine.uiowa.edu/cme/clia/modules.asp?testID=4#02>.

  • Krantz, Alexander. "Residency & Fellowship Programs | Education." Department of Pathology & Cell Biology | CUMC. Columbia University of Physicians and Surgeons. Web. 12 Sept. 2010. <http://pathology.columbia.edu/education/residency>.

  • Harmening., Denise, Clinical Hematology and Fundamentals of Hemostasis, 3rd edition, pp. 593-599.

  • Turgeon, Mary Louise, Clinical Hematology - Theories and Procedures, 3rd edition, pp373, 376-382.

  • Rodak, Bernadette, Diagnostic Hematology, 1st edition, p.605-606.

  • Coulter STKS Operating Manual

  • McKenzie, Shirlyn, Clinical Laboratory Hematology, 2ndedition,pp 813-829.

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