Analysis of human peripheral blood lymphocytes isolated by counterflow centrifugation-elutriation

Abstract

Human peripheral blood mononuclear cells isolated by Ficoll-Hypaque density gradient centrifugation have been fractionated by counterflow centrifugal elutriation (CCE). Six CCE fractions were obtained and subsequently analyzed as for their content of monocytes, T cells, NK cells and B cells. The various cell types were identified through the expression of specific surface membrane determinants or by cytochemical staining for alpha-naphthyl acid esterase (ANAE). Monocytes were elutriated at the highest counterflow rates whereas the majority of B cells were collected at the lowest counterflow rates. T cells as well as NK cells were mostly concentrated in the intermediate fractions. No differences in the elutriation profile of T cells with the helper-inducer or with the cytotoxic-suppressor surface phenotype were observed. However, the percentages of T cells as determined by surface marker expression decreased with increasing counterflow rates, whereas the percentage of ANAE-positive T cells increased. Yet, T cells recovered at the high counterflow rates had ANAE-reactive organelles larger than those of T cells collected at low counterflow rates. These findings suggest that T cells at different maturational stages could be separated by CCE.