Three human basement membranes, glomerular basement membrane (GBM) from renal cortex, alveolar basement membrane (ABM) from lung parenchyma and trophoblast basement membrane (TBM) from the terminal villi of placenta have been isolated by sieving and sonication techniques. Canine GBM and ABM were also prepared. There were marked differences among the membranes from human tissues. Compared to GBM, TBM had very little collagen but contained high concentrations of charged amino acids. ABM was intermediate in composition between GBM and TBM and contained desmosine and isodesmosine indicative of the presence of elastin. Canine ABM (c-ABM) did not contain desmosine or isodesmosine. In the canine system an antigen was detected in ABM which was not present in GBM. The membrane preparations were analyzed for fibronectin content using a specific antiserum to fibronectin. This glycoprotein could not be detected in GBM whereas it was present in ABM in amounts up to 0.8% and in TBM in amounts as high as 7.2%. All the membranes induced the formation of precipitating antibodies in rabbits. Soluble material obtained from the membranes by alkali extraction, reduction of disulfide bonds, enzymatic digestion with elastase, plasmin or collagenase provided immunologically reactive fragments. These soluble fragments gave reactions of identity among the three basement membranes in immunodiffusion reactions in gels with antisera raised to all three BMs. The finding that plasmin digests basement membranes suggests that it may play a role in connective tissue remodeling. The fact that elastase degrades basement membranes provides an endogenous system for injury which may be triggered by infections.