Isolation of human T and B lymphocytes by E-rosette gradient centrifugation. Characterization of the isolated subpopulations

Abstract

Some methodological aspects of E-rosette gradient centrifugation for separation of human B and T lymphocytes are described. Treatment of sheep red blood cells (SRBC) with 2-aminoethylisothiouronium bromide (AET) accelerates and enhances E-rosette formation providing an effective and time saving method which takes less than 2 h to perform. Depletion of E-rosette forming cells (E-RFC) is almost complete, leaving less than 1% of the initial E-RFC at the interphase layer. By use of the lymphocyte donor's autologous serum containing naturally occurring anti-SRBC antibody and complement the SRBC in the rosetted T lymphocyte fraction are easily and completely lysed without damage to the lymphocytes. This procedure is shown to have little effect on the properties of the isolated T cells. The isolated lymphocyte subpopulations have been characterized by tests for lymphocyte and monocyte markers.