[Immunocytochemical determination of membrane-bound IgG using physiologically aged and experimentally aged...

The expression of IgG receptor sites at the band 3 protein is important for the recognition and elimination of aged and experimentally altered erythrocytes. Membrane bound IgG was detected in different erythrocyte preparations and microvesicles by means of electron microscopic procedures (protein A-gold-, protein A-gold-silver- and anti-ferritin-sandwich-technique) and light microscopic procedures (immunofluorescence). Physiologically "old", pronase and neuraminidase as well as diamide treated erythrocytes and microvesicles demonstrated significant IgG loading. An increased IgG binding of erythrocytes treated with phenylhydrazine was only evident when higher phenylhydrazine concentrations were used. Both, the alteration of the glycocalyx (conformational changes of the external segment of the glycophorins) and the alteration of the membrane skeleton lead to an unmasking of the IgG receptor site at band 3 proteins (transmembrane effect). The result is an overcritical loading of cells with IgG molecules which initiate the elimination of the erythrocytes by macrophages of the Reticulo-Histiocytic-System.