Flow cytometric differentiation of erythrocytes and leukocytes in dilute whole blood by light scattering.
We have determined differential scattering cross sections of sphered erythrocytes integrated over the solid angle accepted by our detectors at wavelengths varying between 379.5-632.8 nm, employing Ar+-, Kr+-, and HeNe-laser radiation. Integrated differential cross sections for forward and orthogonal light scatter exhibit a pronounced minimum at about lambda = 413 nm, caused by the strong absorption of light by oxyhemoglobin. Experimental data are in good agreement with values calculated by the Mie theory. Simultaneous measurements of forward and orthogonal light scatter using Kr+-laser radiation (lambda = 413.1 nm) allow flow cytometric differentiation of (native) erythrocytes, thrombocytes, and leukocytes as well as subpopulations of leukocytes, i.e., granulocytes, lymphocytes, and monocytes in dilute whole-blood samples. Lysis of red blood cells or staining of white blood cells is not required to obtain absolute leukocyte counts or differential white blood cell counts.
Abteilung Medizinphysik und metrologische Informationstechnik, Physikalisch-Technische Bundesanstalt, Berlin, Germany.
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Neukammer
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Rinneberg
D001773:Blood Cells / Q000166:cytology; D002469:Cell Separation / Q000295:instrumentation / Q000379:methods; D004906:Erythrocyte Count; D004912:Erythrocytes / Q000166:cytology*; D005434:Flow Cytometry / Q000295:instrumentation / Q000379:methods*; D006801:Humans; D007958:Leukocyte Count; D007962:Leukocytes / Q000166:cytology*; D008027:Light; D012542:Scattering, Radiation