Precipitability and composition of HBsAg-anti-HBs immune complexes formed in the presence of complement. A model of circulating immune complex analysis.
Immune complexes (IC) of partially purified HBsAg and human anti-HBs were prepared at different antigen/antibody ratios in the presence of complement in normal human serum (NHS), and under conditions not allowing complement activation in buffers or in NHS containing 10 mM EDTA (NHS-EDTA). Commercial preparations of the radiolabelled antigen and antibody were used. IC formed in NHS were not significantly precipitated even after incubation for 24 h at 4 degrees C, whereas a typical precipitation curve was observed with complexes formed in the absence of complement. Thus, complement activation was found to markedly and permanently inhibit precipitability of HBsAg-anti-HBs immune complexes (HBsAg-IC). HBsAg-IC were precipitated from sera with 3.5% polyethylene glycol (PEG), boiled in sodium dodecyl sulphate (SDS)-urea buffer, and analysed by SDS-polyacrylamide slab gel electrophoresis (SDS-PAGE). With complexes formed in the presence of complement, about one-sixth of the antibody activity was found in high molecular weight fractions corresponding in size to IgG oligomers. By contrast, with complexes formed without complement, no significant amount of antibody was found in these fractions. With blotting technique and radiolabelled anti-human-C3 antibody, it was demonstrated that anti-HBs was covalently bound to C3b fragments in IC formed in the presence of complement and was in the high molecular weight fractions.
D000818:Animals; D000936:Antigen-Antibody Complex / Q000032:analysis* / Q000276:immunology; D000937:Antigen-Antibody Reactions; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D003179:Complement C3b / Q000378:metabolism; D003165:Complement System Proteins / Q000502:physiology; D004591:Electrophoresis, Polyacrylamide Gel; D006168:Guinea Pigs; D006510:Hepatitis B Antibodies / Q000276:immunology*; D006514:Hepatitis B Surface Antigens / Q000276:immunology*; D006801:Humans; D007158:Immunologic Techniques; D008954:Models, Biological; D008970:Molecular Weight; D011092:Polyethylene Glycols
D000936:Antigen-Antibody Complex; D006510:Hepatitis B Antibodies; D006514:Hepatitis B Surface Antigens; D011092:Polyethylene Glycols; D003179:Complement C3b; D003165:Complement System Proteins