Isolation of human mononuclear cell subsets by counterflow centrifugal elutriation (CCE). I. Characterization of B-lymphocyte-, T-lymphocyte-, and monocyte-enriched fractions by flow cytometric analysis.
Rapid separation of large numbers of human peripheral blood mononuclear cells into fractions enriched for B lymphocytes, T lymphocytes, or monocytes was accomplished by counterflow centrifugal elutriation (CCE). The first fraction contained 98% of the platelets. Ten additional fractions containing subpopulations of mononuclear cells were collected by sequential increases in the flow rate while maintaining a constant centrifuge speed. Analysis of the fractions using monoclonal antibodies revealed that fraction 2, which was free of esterase-positive monocytes, was highly enriched for B cells. T lymphocytes (OKT3+) were the predominant cell type found in fraction 4. No enrichment for T-lymphocyte-helper (OKT4+) or -suppressor (OKT8+) subpopulations was observed in the lymphocyte containing fractions. Three fractions (7-9), highly enriched for esterase-positive cells, were predominantly OKM1+ monocytes with no evidence of selective separation of monocyte subpopulations. Thus, cell fractions enriched for B cells, T cells, and monocytes could be obtained, by utilizing CCE, in large enough quantities to enable analysis of their functional properties. Of particular interest was the ability to separate small, resting B lymphocytes from monocytes.
D000911:Antibodies, Monoclonal / Q000276:immunology; D000954:Antigens, Surface / Q000276:immunology; D001402:B-Lymphocytes / Q000145:classification* / Q000276:immunology / Q000378:metabolism; D002469:Cell Separation / Q000379:methods*; D002499:Centrifugation, Density Gradient; D004950:Esterases; D005434:Flow Cytometry; D006801:Humans; D009000:Monocytes / Q000145:classification* / Q000276:immunology / Q000378:metabolism; D011947:Receptors, Antigen, B-Cell / Q000032:analysis; D011961:Receptors, Fc / Q000032:analysis; D017452:Receptors, IgG; D013601:T-Lymphocytes / Q000145:classification* / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D011947:Receptors, Antigen, B-Cell; D011961:Receptors, Fc; D017452:Receptors, IgG; D004950:Esterases