Purification of human granulocytes by centrifugal elutriation and measurement of transmembrane potential

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Abstract
Viable cell samples containing 93% pure granulocytes were obtained from human blood using the techniques of dextran sedimentation followed by centrifugal elutriation. The resting transmembrane potential (Em) of human granulocytes was estimated using the fluorescent lipophilic cation, Di-S-C3(5), from the null point for potassium—i.e., the external K concentration at which there is no change in Em in response to valinomycin (a K ionophore). The Em of human granulocytes, as calculated from the Nernst potential for K at the null point, is approximately – 100 mV. Data indicate that this large transmembrane potential is due in part to the presence of an electrogenic Na-K pump in human granulocytes which is stimulated by external potassium and inhibited by ouabain.