Purification of soluble immune complexes from serum using polymethylmetacrylate beads coated with conglutinin or C1q. Application to the analysis of the components of in vitro formed immune complexes and of immune complexes occurring in vivo during leishmaniasis.
A procedure for the isolation of immune complexes from human sera has been developed. Two steps are involved: (1) lipid-free serum is precipitated by polyethylene glycol; (2) the solubilized precipitate is absorbed on a column of polymethylmethacrylate beads coated with conglutinin (K) or C1q; the column is washed, the complexes are then eluted, using 0.02 M EDTA (for K column) or 0.5 M NaCl (for C1q column). This procedure permitted the purification and the characterization of soluble 125I-BSA-anti-BSA, 125 I-tetanus toxoid-anti-tetanus toxoid, and 125-I-HBsAg-anti-HBsAg complexes made in vitro in the presence of fresh human serum. The isolated complexes were shown to contain antigen, antibody, C1q, C1r, C1s and C3. When normal human serum was submitted to such a procedure, no detectable amount of protein was present in the final eluted fraction. Immune complexes formed in vivo were also purified by conglutinin column from the serum of a patient with disseminated leishmaniasis. The isolated material was found to contain IgM, IgG, C1q, C1r, C1s, C3c and C3d. The purified complexes dissociated at acid pH were found to contain anti-IgG and anti-leishmania antibodies.
D000328:Adult; D000936:Antigen-Antibody Complex; D000941:Antigens / Q000032:analysis; D003172:Complement C1 / Q000032:analysis; D003176:Complement C3 / Q000032:analysis; D006801:Humans; D007122:Immunoelectrophoresis; D007074:Immunoglobulin G / Q000032:analysis; D007136:Immunoglobulins / Q000032:analysis; D007898:Leishmaniasis, Visceral / Q000276:immunology*; D008722:Methods; D011109:Polymethacrylic Acids; D012995:Solubility
D000936:Antigen-Antibody Complex; D000941:Antigens; D003172:Complement C1; D003176:Complement C3; D007074:Immunoglobulin G; D007136:Immunoglobulins; D011109:Polymethacrylic Acids