Determination of Number and Size of Particles by Electrical Gating: Blood Cells

¹ From the National Institutes of Health, Bethesda, Maryland

An instrument is described which applies principles of electrical gating to the problem of particle counting. Cells are suspended in an electrically conducting medium which flows through a small aperture. The relatively nonconductive cells cause a voltage drop as they pass through the apperture. The resulting pulses are amplified and appear on an oscilloscope. Those exceeding a given threshold level are recorded on a decade counter. A 1/50,000 dilution of whole blood in normal saline results in the enumeration of about 50,000 red cells in 0.5 cc. The instrument count must be corrected upward because of the occasional passage through the aperture of two or more cells simultaneously. This coincidence follows the Poisson distribution and can be corrected for by a calibration curve. The instrument demonstrated excellent reproducibility. Instrument counts, corrected by a dilution calibration curve, were consistently lower than the mean of multiple hemocytometer counts. However, the hemocytometer counts performed in this laboratory were found to be subject to a systematic error arising from technique of hemocytometer loading. The error is believed to result from planar concentration of cells flowing into the chamber. Since a relationship exists between the pulse height and particle size, a type of size distribution curve may be obtained by taking successive counts at increasing threshold heights. By this means, the number of pulses of a given increment of height may be obtained which when plotted against threshold level results essentially in a first derivative of the original curve. Thus one obtains a pulse height distribution curve related to the size distribution of the particles. These curves have been found quite similar to Price-Jones curves on a number of blood samples of various origin. Preliminary studies in white cell counting are presented. Saponin has been used to effectively hemolyze and stromatolyze erythrocytes apparently without serious damage to leucocytes. Instrumental and hemocytometer white cell counts were found to be in reasonably good agreement.

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