A procedure to isolate soluble IgG-containing immune complexes from serum was developed, which combined Sephadex G-150 filtration with affinity chromatography on Staphylococcal protein A-Sepharose. The effectiveness of the method was shown by isolating model immune complexes of 125I bovine plasma albumin (BPA) and antibody to BPA from serum. Functionally intact antibody and antigen were recovered from these complexes by chromatography on Sephadex G-150 at low pH and the degree of purification was demonstrated by NaDod.SO4 polyacrylamide slab gel electrophoresis (PAGE) and autoradiography of the immune complexes and their components.

Application of these methods to a Gross virus-induced rat lymphoma model tested their potential for characterizing naturally occurring circulating immune complexes (CIC). Sera from these animals were shown by Raji cell immunofluorescence to contain high concentrations of CIC 15 to 21 days after tumor inoculation. CIC could be isolated from these sera by the procedures developed for model complexes. Analysis by PAGE indicated that they contained primarily a protein of apparent m.w. 79,000 complexed with IgG. This putative antigen component of CIC (79,000 daltons) reacted with a rabbit antiserum directed against the murine leukemia virus (MuLV) envelope glycoprotein, gp70. Techniques using protein A affinity chromatography may also be applicable to characterization of both antigen and antibody components of CIC in human diseases including cancer.

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