Male Sprague-Dawley rats developed acute lung and renal injury after administration of heterologous anti-lung antibody. Both organs demonstrated an increase in protein permeability after antibody binding to basement membrane (BM) antigens (lung permeability index 0.342 +/- 0.009 versus control 0.214 +/- 0.011: p less than 0.05. Urinary protein excretion 5.112 +/- 0.899 mg/hour versus control 0.402 +/- 0.008 mg/hour: p less than 0.01). The threshold value for the development of lung injury was 27.2 +/- 4.8 micrograms of antibody globulin/g of tissue (micrograms/gm). Immunoblot analysis probing with the anti-lung antibody revealed at least one common antigenic determinant (82 to 84 kilodaltons) bound within collagenase-solubilized pulmonary and glomerular BMs. Increasing doses of antibody produced hemorrhagic pneumonitis and diffuse alveolar damage. Immunofluorescence microscopy confirmed linear alveolar and glomerular BM antibody binding. Immunogold electron microscopy allowed precise identification, in intense linear patterns, of BM binding sites within lung and glomeruli. Functional lung injury was prevented by either leukocyte-depletion or complement-depletion (lung permeability index antibody-treated, complement-depleted 0.235 +/- 0.034: both p greater than 0.05 compared with controls). Injury mediation in this acute humoral model of lung damage is both complement- and leukocyte-dependent, as previously described for the renal component of heterologous anti-glomerular BM antibody-induced inflammatory disease.

Post-embedding immunogold electron microscopy (IAuEM) techniques utilizing low-temperature embedding in Lowicryl K4m and LR White resin were used to localize the binding sites of two sheep anti-rat glomerular basement membrane polyclonal antibodies (P1 and P2) and a sheep anti-rat lung antigen antibody. P1 localization was bilaminar in a linear pattern along the subepithelial and particularly the subendothelial aspect of the glomerular basement membrane. P2 was bound diffusely throughout the lamina densa, even at supramaximal doses. The anti-lung antibody bound in an interrupted linear pattern throughout the lung basement membrane (alveolar and capillary), and showed an intense, diffuse binding to the glomerular basement membrane. IAuEM allowed definition of the precise basement membrane binding sites of these polyclonal reagents.