LYMPHOCYTE SUBPOPULATIONS IN PERIPHERAL BLOOD OF HEALTHY PERSONS.

Lymphocytes were isolated at 99% purity from peripheral blood of healthy persons by defibrination, gelatine sedimentation, treatment with carbonyl iron powder and centrifugation on Ficoll-Isopaque. Subpopulations were identified by three surface markers: cells forming rosettes with sheep red blood cells (SRBC) (E-binding lymphocytes) as a measure of T lymphocytes: lymphocyte with surface immunoglobulin identified by indirect immunofluorescence (B lymphocytes): lymphocytes with receptors for C3 observed by the rosette method using SRRC treated with rabbit antiserum and human complement (EAC-binding lymphocytes). The yield of lymphocytes after purification varied from 15 to 65%. No selection of lymphocytes was observed either by counting immunoglobulin-bearing and EAC-binding lymphocytes in whole blood and in purified cells from the same sample. or histatistical analysis of lymphocytes in subpopulations as a function of the yields from twenty-six experiments. In the absence of selection during purification the total numbers of T and B lymphocyte could be calculated from the percentages and the total numbers of lymphocytes. Our normal values are close to those reported using other non-selective methods of purification. When lymphocytes were simultaneously stained for immunoglobulin and rosetted with EAC, cells bearing either or both markers were found. In total, 27- 35% cells were identified by these markers. Since about 70% of the cells were E- binding, practically all lymphocytes could be identified. A small overlap between F-binding and immunoglobuIin-bearing EAC-binding lymphocytes may occur. Either the IgM or the IgG-containing fractions obtained after fractionation of rabbit anti-SRBC serum on Sephadex G-200 could be used for sensitization of SRBC with complement. Formation of rosettes was not prevented by pretreating the lymphocytes with aggregated IgG, while rosettes formed with EA prepared by high concentrations of IgG antibody (Fe-binding lymphocytes) were abolished. It is concluded that rosettes formed with IgG-FA( (or whole serum FAC) usingdiluted antiserum identify complement-reactive lymphocytes and are not caused by synergism with Fc receptors. When SRBC were sensitized with varying dilutions of whole antiserum or in IgG fraction identical plateaus for the percentages of LAC- binding lymphocytes were found. Subagglutinating concentrations of the IgM fraction was insufficient to reach the plateau and also consistently resulted in lower values for EAC-binding lymphocyte. [ABSTRACT FROM AUTHOR]