It is accepted that the main determinant of glomerular injury in experimental nephrotoxic
nephritis is the administered dose of anti-glomerular basement membrane (GBM) antibody. However, there are other factors that can enhance the severity of such injury including small doses of bacterial
lipopolysaccharide (LPS). In the present study, we have assessed whether preparations of
anti-GBM antibody contaminated with different concentrations of
endotoxin could influence the severity of glomerular injury in the heterologous phase of nephrotoxic
nephritis. We have also examined the efficacy of different laboratory methods to isolate an
endotoxin-free
anti-GBM antibody, and to purify
anti-GBM antibody preparations from
endotoxin. Preparations of
anti-GBM antibody (nephrotoxic
globulin) isolated from nephrotoxic serum by the
sodium sulphate precipitation method contained variable concentrations of
endotoxin. Administration of these preparations in equal doses into clean rats, which had no established
acute phase response, markedly aggravated the severity of glomerular injury. However, preparations contained less than 50 pg/ml of
endotoxin appeared to have no significant effect on such injury. Furthermore, isolation of
anti-GBM antibody from nephrotoxic serum by affinity chromatography, using Staphylococcus
protein-A column, proved to be a reliable method not only for the isolation of an
IgG (nephrotoxic antibody) free from other serum contaminants, but also for purification of
endotoxin contaminated preparations of
anti-GBM antibody. These observations have practical implications in studying models of
nephritis as our results show that the glomerular injury, which is usually considered to be a sole function of the mass of antibody bound to GBM, is profoundly influenced by minor
endotoxin contamination of the
anti-GBM antibody.