Monocyte purification with counterflow centrifugation monitored by continuous flow cytometry

Abstract

Continuous monitoring of cell light scatter during counterflow centrifugation of a mononuclear cell suspension allows counting and size recognition of the cell types elutriated. With this method an optimal separation point between monocytes and lymphocytes, determined for each individual donor, may be established. With a constant flow of 15 ml/min this separation point is found at centrifugal velocities ranging from 2348 to 2444 rpm (n = 10). From 50 ml venous blood, 84.1% +/- 4.1% (15.7 +/- 8.6 x 10(6)) of all elutriated monocytes, with a purity of 92.4% +/- 1.4%, is collected in a volume of 50 +/- 1 ml. In the same run, 92% +/- 4.3% of the lymphocytes is gathered in one fraction with a purity of 98.9% +/- 0.7%. After counterflow centrifugation, 91.6 +/- 10.5% of the cells loaded is recovered; viability exceeds 98%.