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Fractionation of Human Blood Lymphocytes on Helix pomatia A Haemagglutinin Coupled to Sepharose Beads

1976; Wiley; Volume: 5; Issue: s5 Linguagem: Inglês

10.1111/j.1365-3083.1976.tb03855.x

1365-3083

Ulla Hellström, Sten Hammarström, M.‐L. DILLNER, Hedvig Perlmann, Peter Perlmann,

Parasites and Host Interactions

Abstract. Treatment of human blood lymphocytes with neuraminidase has previausly been shown to uncover receptors for the A haemagglutinin of the snail Helix pomatia (HP). Neuraminidase‐treated lymphocytes were now fractionated on columns charged with large Sepharose particles to which HP had been coupled covalently. HP‐receptor negative (HP ‐ ) lymphocytes passed the columns while HP‐receptor positive (HP+) lymphocytes were retained. The latter cells were eluted by addition of the competitive hapten N‐acetyl‐D‐galactosamine (D‐GalNAc). The total yield of cells recovered after fractionation was 60‐80% Surface marker studies indicated that there was no selective loss of any of the major lymphocyte subpopulations. The fraction that passed the columns (fraction I) consisted of approximately 10% of all lymphocytes. It contained ˜ 1% HP+ cells and ˜3% of all lymphocytes forming rosettes with sheep erythrocytes (E+ cells) present before fractionation. 50‐55% of the lymphocytes in this fraction had surface‐bound immunoglobulin (SIg+ cells) and complement receptors (EAC+ cells). Of the SIg+ cells, ˜60% were true B cells while the remaining 40% had IgG adsorbed to their surface. The majority of the B cells were recovered in this fraction. The lymphocytes of this fraction responded poorly to T‐cell mitogen but had an enhanced K‐cell activity to chicken erythrocytes. Elution of the cells retained on the column with 0.1 mg/ml D‐GalNAc gave a fraction II, consisting of, ˜15% of all lymphocytes. This fraction had a mixed composition. The majority of the cells (˜45%) were recovered by subsequent elution with 1.0 mg/ml D‐GalNAc. This fraction III was strongly enriched with HP+ and E+ cells (T cells). About 10% of the HP+ cells in this fraction were SIg+. However, on the majority of these cells this surface‐bound immunoglobulin was probably externally adsorbed IgG. These HP+‐SIg+ cells were also EAC+ and had Fc receptors, as shown by rosette formation with IgG‐coated bovine erythrocytes. The lymphocytes of fraction III responded most strongly to T‐cell mitogen while their K‐cell activity was weak.