Proliferative and Cytolytic Reactivities of Human Lymphocytes Fractionated on Wheat Germ Agglutinin‐Sepharose Columns before or after Alloactivation in Mixed Lymphocyte Culture

Abstract

Human T-lymphocyte preparations from peripheral blood with either high- or low-avidity receptors for wheat germ agglutinin (WGA) were obtained by fractioned on WGA--Sepharose columns. Both fractions contained progenitors of alloreactive T cells, proliferating in mixed lymphocyte culture (MLC) and acting as effector cells in cell-mediated lympholysis (CML). Proliferation and CML activity of the two fractions were equal and similar to those of the unfractionated cells. However, when the lymphocytes were fractionated after 5 days' MLC, most of the proliferating and cytolytic cells were found in the lymphocyte fraction enriched in cells with high-avidity receptors for WGA. The reactivity of the fractions was correlated to their content of blast-transformed cells. The binding of MLC-activated lymphocytes to WGA was specific, since it was inhibited by the competitive hapten D-GlcNAc and no cells were retained on control columns charged with human serum albumin-Sepharose. B cells and monocytic cells were enriched in the fraction with low-avidity receptors for WGA. As indicated by experiments in which cells were mixed in different proportions, the low MLC-CML activity of the lymphocytes in the fraction with low-avidity WGA receptors was not caused by suppressor cells present in that fraction.

Analysis of the T‐Cell Receptor Vβ Usage in Monozygotic and Dizygotic Twins Living in a Plasmodium falciparum Endemic Area in West Africa

“…Cells forming E-rosettes were separated from non-E-rosetting cells by Ficoll-Isopaque density centrifugation. Sheep erythrocytes attached to T lymphocytes were eliminated by osmotic lysis [17]. Highly purified T cells were separated into CD4 + and CD8 + cells by negative depletion using magnetic beads coupled with anti-CD4 or anti-CD8 antibodies (Dynal AS, Sköyen, Oslo, Norway).…”

Enhancement of Human B‐Cell Proliferation by a Monoclonal Antibody to CD43

“…type VI. Sigma) treated sheep erythrocytes [18]. The non-rosetting cell population was further depleted of monocytes by adherence in plastic tissue culture flasks for 1 h at 37 C in Hepes-buflered RPMI-1640 containing 50"'.. heat-inactivated human normal AB' scrum (HNS).…”

Separation of Human B Lymphocytes on Helix pomatia A Haemagglutinin into Two Major Fractions Differing in Responsiveness to T‐dependent Mitogen (Pokeweed Mitogen) or Antigen (Tetanus Toxoid)

“…The cells were washed with Tris-buffered Hank's solution (TH)+0.2% human serum albumin (HSA). They were rosetted with neuraminidase-treated sheep erythrocytes in undiluted fetal calf serum (FCS) (heat inactivated for 60 min, 56°C) at 4°C overnight as described in detail earlier [24].…”

“…The rosette-forming T cells in the Ficoll-Paque pellet were resuspended and dissoeiated by incubation at 37°C for 15 min, with frequent Vortex mixing. The sheep erythrocytes were lysed with distilled water [24]. The T cells were resuspended in RPMI 1640 buffered with 10 mM Hepes (Biocult Laboratories, Paisley, Scotland, UK) and supplemented with 20% FCS, 2 mM L-glutamine, penicillin 100 IU/ml, and streptomycin 100 fig/ml (complete TCM).…”