Membrane distribution and adsorptive endocytosis by C3b receptors on human polymorphonuclear leukocytes.

C3b receptors on human polymorphonuclear leukocytes (PMN) were nonrandomly distributed in small clusters on the plasma membranes of these cells when assessed by indirect immunofluorescence at 0 degree C using monospecific rabbit Fab' or F(ab')2 anti-C3b receptor and tetramethylrhodamine isothiocyanate (TRITC)-conjugated goat IgG anti-F(ab')2. When PMN were incubated with the bivalent anti-C3b receptor at 37 rather than at 0 degree C, almost no immunofluorescence was observed, which indicates that the C3b receptor-F(ab')2 complexes had been rendered inaccessible to TRITC-IgG anti-F(ab')2. Endocytosis of the anti-C3b receptor ligand was quantitated by measuring the binding 131I-IgG anti-F(ab')2 by PMN that had previously taken up 125I-F(ab')2 anti-C3b receptor at 0 and at 37 degree C, respectively. There was a constant 2: 1 molar ratio of anti-F(ab')2 to anti-C3b receptor with PMN that had been incubated with the first antibody at 0 degree C. In contrast, when increments of F(ab')2 anti-C3b receptor were taken up by the cells at 37 degree C, there was a dose-related decline in this molar ratio to a minimum of 0.2 molecules of anti-F(ab')2 anti-F(ab')2 bound per molecule of PMN-associated anti-C3b receptor. 125I-F(ab')2 anti-C3b receptor taken up by PMN at 37 degree C was also inaccessible to release by proteolytic treatment of the cells with pronase. The rate of endocytosis of 125I-F(ab')2 anti-C3b receptor was rapid as the PMN-bound antibody fragment became inaccessible to 131I-IgG anti-F(ab')2 within 10 min during incubation of the cells at 37 degree C. In contrast to these findings, 125I-Fab' anti-C3b receptor that was taken up by PMN at 37 degree C remained accessible to both 131I-IgG anti-F(ab')2 and to proteolytic release by pronase, which suggests that monovalent interaction of ligand with C3b receptors was not sufficient for induction of endocytosis. The requirement for multivalency was also demonstrated using the C3b-OR, the normal ligand for the C3b receptor. 125I-C3b-OR was specifically bound by PMN but remained on the cell receptor. 125I-C3b-OR was specifically bound by PMN but remained on the cell surface, as determined by its accessibility to pronase, unless it was cross-linked with F(ab')2 anti-C3. Although C3b receptors on PMN do not mediate internalization of adsorptive pinocytosis of soluble ligand indicates their potential for the clearance of C3b-bearing immune complexes without recruitment of other cell surface receptors.