Electron Microscopic Studies of Experimental Nephritis With Ferritin-Conjugated Antibody

Abstract

The acute nephritis that rapidly follows the injection of antikidney serum is the result of a specific reaction between antigens localized in the glomeruli and antibodies contained in the nephrotoxic sera. The antibodies, demonstrated by means of the fluorescence technique, were found in greatest concentration apparently in or on the glomerular basement membrane of nephritic animals and were noted to remain there for many months (1). The precise localization of nephrotoxie globulin in the glomeruli, however, was dif]fieult to establish owing in part to the limited resolution of the ultraviolet light microscope and in part to the relatively thick sections employed.This paper reports the use of ferritin-conjugated antibody globulin (2) to identify the sites of antigen-antibody interaction by electron microscopy. This technique has been applied to the localization of viral and bacterial antigens (3-6). Preliminary studies from this laboratory on the application of ferritinconjugated antibody to the study of experimental glomerulonephritis have been described elsewhere (7). Materials and MethodsThe sera used to produce nephritis were the same rabbit antisera to rat kidney employed in groups I and 3 referred to in reference 1. Long-Evans rats, varying in age from 2 to 3 months, were injected intravenously on 2 or 3 successive days with a high titered antikidney serum which produced proteinuria and renal lesions within 24 hours. The animals were killed 1 week later.

The Histogenesis of Basement Membranes

“…In fact the cells may lose the ability to synthesize, which may account for the lack of renal tubular regeneration when there is extensive damage to both the epithelium and its basement membrane. Other observations (1,3) have shown that the endoplasmic reticulum of the normal glomerular epithelium may occasionally contain basement membrane material.…”

“…A common cross-reacting antigen, for instance, has been observed in studies of tissue-specific antigens and has been attributed to a common vascular antigen, presumably originating in the basement membranes (22). It has been demonstrated that renal glomerular basement membranes are antigenic when injected in heterologous species (8,23); the antibody elicited is capable of causing severe anatomical and functional derangement of the renal glomeruli when injected into the original species (1,8,(23)(24)(25). When the mechanism of this experimental glomerulonephritis was investigated using fluorescent antibody techniques, basement membranes around epithelia and within vessels fluoresced specifically as did reticulin (25).…”

Comparison of Reactions of Antibodies to Rat Collagen and to Rat Kidney in the Basement Membranes of Rat Renal Glomeruli

“…This fluorescence differs from the irregular granular appearance of the capillary wall which is seen when antigen-antibody complexes are deposited in the kidney, as, for example, in serum sickness nephritis (17). The specific site of reaction of ferritinconjugated nephrotoxic antibody has been demonstrated by electron microscopy within the renal glomerular basement membranes (18)(19)(20).…”

Electron Microscopic Studies of Experimental Nephritis With Ferritin-Conjugated Antibody

“…The observations reported here confirm their data. The immuno-ferritin technique, employed in the study of nephrotoxic nephritis in rats (6) has been applied to the investigation of the glomerulonephritis produced by the method of Dixon and his associates.…”

“…The tagging of the renal tissue with ferritin-conjugated antibody to BSA was blocked by first treating the tissue with unlabeled antiserum according to the technique already described (6).…”