Lockwood Cm

A mouse monoclonal antibody (P1) to the autoantigenic component of human glomerular basement membrane (GBM) was used to study the immunochemistry and tissue distribution of the Goodpasture antigen and the specificity of the human autoimmune response in Goodpasture's syndrome (anti-GBM disease). In solid phase assays, monoclonal antibody P1 bound to collagenase-solubilized human GBM (the ligand used in assays for human autoantibody), but not to other biochemically defined components of basement membrane. On Western blotting, P1 bound to the same 6 bands in solubilized GBM (between 26 and 58 kilodaltons with major bands at 26 and 54 kilodaltons) that were recognized by sera from all 42 patients studied with anti-GBM disease. Preincubation with sera from 8/8 patients blocked the subsequent binding of P1 from 83 to 89% on densitometer scanning of the Western blot; and preincubation with P1 blocked the binding of sera from 6/6 patients from 58 to 89%. Indirect immunofluorescence and immunoperoxidase studies revealed that the pattern of binding of P1 was identical to that of antibody eluted from the kidneys of a patient with Goodpasture's syndrome; there was linear binding to GBM, Bowman's capsule, and distal tubular basement membrane. In addition, P1 bound to basement membranes in lung and choroid plexus, and to membranes of the lens capsule, choroid, and retina of the eye and cochlea, but not to other organs studied. It is concluded that there is a single major autoantigenic component of human GBM (the Goodpasture antigen), which is present on fragments of different molecular weight in the collagenase digest. This antigen is distributed throughout well-defined basement membranes known to be involved in both Goodpasture's and Alport's syndromes. Human anti-GBM antibodies bind to the same (or closely related) determinants which are recognized by P1, demonstrating that the autoimmune response in Goodpasture's syndrome is of highly restricted specificity.

We describe a sensitive radio-immunoassay for auto-antibodies to glomerular basement membrane (GBM). The assay discriminates patients with glomerulonephritis associated with circulating auto-antibodies to glomerular basement membrane from those with other forms of nephritis. Monitoring of antibody levels in sequential serial serum samples enables drug therapy to be tailored precisely to control the aberrant immune response, a situation unique in the management of glomerulonephritis.

Hereditary nephritis (HN) is associated with antigenically abnormal glomerular basement membrane (GBM) manifest by reduced or absent binding of MCA-P1, a mouse monoclonal antibody which recognizes Goodpasture antigen. In the present studies, immunoblotting has been used to analyze antigenic and biochemical composition of renal tissue from patients with HN in whom binding of MCA-P1 could not be demonstrated by indirect immunofluorescence (IIF). Pooled normal collagenase-solubilized GBM (CS-GBM) and CS-GBM from three patients with either end-stage renal failure (ESK1-3) or HN (HNK1-3), were examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis gel and, after transfer by Western blotting to nitrocellulose, reacted with sera from six patients with anti-GBM disease (GPS1-6) or anti-GBM antibody containing sera from three transplanted HN patients (HNS1-3), different from those patients with HN contributing HNK1-3. We found that GPS1-6 and HNS1-2 recognized the same six major bands in CS-GBM and ESK1-3 (between 54 and 24 kilodalton (kd) but only three bands (48, 42 and 24 kd) in HNK1-3. HNS3 only bound strongly to bands of 54 and 26 kd in CS-GBM or ESK1-3 and not all to HNK1-3. Immunoblotting studies of HNK1-3 have shown a partial rather than absolute loss of Goodpasture antigenicity (54, 28 and 26 kd bands). Studies with HNS1-3 suggest heterogeneity of antibody responses to allografted kidneys between patients with HN; HNS-3 showed restricted antibody specificity with recognition in CS-GBM of some bands antigenically absent from HN kidney. The abnormality in HN kidneys appears closely related to, but distinct from, the Goodpasture determinant and the inherited defect in HN may involve an essential modifying enzyme.