Characterization of nephritogenic IgA immune complexes separated by polyethylene glycol

The size of IgA immune complexes (IgA-IC) plays an important role in the pathogenesis of experimental IgA nephropathy. The ability of different concentrations (3.5%, 5% and 7% w/v) of polyethylene glycol (PEG) to selectively precipitate IgA-IC with defined size, was examined using convalently cross-linked 125I-radiolabelled IgA-IC. These complexes were prepared with purified IgA anti-dinitrophenyl (DNP) antibodies and bis-DNP-pimelic acid ester. Size analysis of IgA-IC by gradient polyacrylamide gel electrophoresis (GPAGE) revealed a composition of 33% large-size (> 2 x 106 mw), 39% intermediate-size (4-20 x 105 mw), and 28% mixture of dimer and monomer IgA. The standard 3.5% PEG precipitated less than 7% of large- and intermediate-size IgA-IC. In contrast, 5% and 7% PEG precipitated 40% and 100% of large-size complexes, respectively. The 7% PEG was also effective in precipitating (40%) of the intermediate-size complexes. The correlation between PEG concentration and size of the precipitated IgA-IC was further confirmed by GPAGE and quantitative autoradiography of resolubilized PEG precipitates of discrete size IgA-IC obtained by gel filtration. They conclude the standard 3.5% PEG is inappropriate for precipitation of IgA-IC and recommend the use of 7% PEG for the detection of intermediate- and large-size IgA-IC