Preliminary experiments on serum from 12 patients with membranoproliferative glomerulonephritis (MPGN) have identified material sedimenting at 7S which binds to solid phase (sp) Clq but not to fluid phase Clq. Similar observations have been made by others using SLE serum. The 7S material reacts with anti-human IgG and studies of others as well as our own have provided compelling evidence that it is antibody to cryptic determinants on Clq revealed when Clq is deposited on polystyrene or on an immune precipitate. The proposed studies will provide further evidence that 7S sp Cla-binding material is anti-sp Clq and will investigate the consequences of such an antibody in vivo. To identify the sp Clq-binding material as antibody, F(ab')2 will be made from that eluted from Clq bound to Sepharose 4B-IgG. Binding of this F(ab')2 to sp Clq on polystyrene will be detected by an ELISA assay employing peroxidase labeled anti-human F(ab')2. To determine whether it will bind to Clq deposited on erythrocytes, sheep EAClq will be reacted with IgG isolated from MPGN serum or NHS as a control and then with 125I anti-human IgG to detect IgG. Binding of anti-sp Clq to EAClq may cause a """"""""multiplier effect"""""""" on complement activation by making more Fc available to activate the classical pathway. This will be assessed by determining whether addition of this autoantibody to EAClq will enhance cell lysis in the presence of complement. Evidence for an additional """"""""multiplier effect"""""""" on immune complex size will be sought by isolating greater than 19S Clq-binding material by sieve chromatography from SLE serum, absorbing it to Sepharose 4B-IgG-Clq and eluting with acid. Identification of 7S anti-sp Clq in the acid eluate will constitute evidence that this autoantibody contributes to the immune complex size. Attempts will be made to develop a rapid method for measurement of anti-sp Clq based on the fact that reduction and alkylation of IgG destroys the Fc but spares the Fab. Fab binding to sp Clq will be measured by ELISA. Using this method, anti-sp Clq will be measured in stored serial serum samples from patients with MPGN and other nephritides and the results correlated with clinical course, complement profile and the results of renal biopsies. Finally, frozen sections of renal biopsies representative of a number of glomerulonephritides will be studied with fluorescein labeled anti-sp Clq isolated from MPGN serum to determine whether anti-sp Clq is a part of the glomerular immune deposits.
| Strife, C F; Leahy, A E; West, C D (1989) Antibody to a cryptic, solid phase C1Q antigen in membranoproliferative nephritis. Kidney Int 35:836-42 |
