Helix pomatia agglutinin (HpA) affinity chromatography: the isolation of pure B and T cell populations and their use for the routine HLA-DR (Ia) serology.
The anti-A1 agglutinin (HpA) of the albumin gland of the snail Helix pomatia binds specifically to neuraminidase-treated T lymphocytes. After its immobilization on Sepharose-6MB an affinity matrix is obtained which is able to separate T and B lymphocytes. In 40 independent experiments enriched T and B cell populations were isolated either by this HpA affinity technique or by E-rosettes incubated with sheep red blood cells for 2 h or 15 h. The results showed that with the HpA affinity matrix within 3 h a highly enriched B cell population is isolated with a purity of 80% and a yield of 65%. Since the separation is done within a short period of time and under gentle conditions the percentage of dead cells is not more than 5%, which is a requirement when the cells are to be used for immunofluorescence studies of surface markers and serological determinations involving microscopical scoring. Typing for HLA-A, B, C antigens and for HLA-DR alloantigens can be done on the same day by cytotoxicity assay. The method also provides a clear-cut distinction between the HLA-A, B, C antibodies and the HLA-DR antibodies.
D000373:Agglutinins / Q000302:isolation & purification; D000818:Animals; D001402:B-Lymphocytes; D002469:Cell Separation; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D006680:HLA Antigens; D006372:Helix, Snails / Q000276:immunology*; D006384:Hemagglutination; D006801:Humans; D005779:Immunodiffusion; D007122:Immunoelectrophoresis; D011817:Rabbits; D012397:Rosette Formation; D013601:T-Lymphocytes
D000373:Agglutinins; D006680:HLA Antigens