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

• Published in: Cellular immunology Vol. 85; no. 2; pp. 373 - 383
• Main Authors: Wahl, Larry M., Katona, Ildy M., Wilder, Ronald L., Winter, Christine C., Haraoui, Boulos, Scher, Irwin, Wahl, Sharon M.
• Format: Journal Article
• Language: English
• Published: San Diego, CA Elsevier Inc 1984; Elsevier
• Subjects: Analysis of the immune response. Humoral and cellular immunity; Antibodies, Monoclonal - immunology; Antigens, Surface - immunology; B-Lymphocytes - classification; B-Lymphocytes - immunology; B-Lymphocytes - metabolism; Biological and medical sciences; Cell Separation - methods; Centrifugation, Density Gradient; Esterases; Flow Cytometry; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Humans; Immunobiology; Monocytes - classification; Monocytes - immunology; Monocytes - metabolism; Receptors, Antigen, B-Cell - analysis; Receptors, Fc - analysis; Receptors, IgG; T-Lymphocytes - classification; T-Lymphocytes - immunology; T-Lymphocytes - metabolism
• ISSN: 0008-8749; 1090-2163
• DOI: 10.1016/0008-8749(84)90251-X

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 predominent 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 predominately 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.