Immunoelectron microscopic localization of immunoglobulins and complement in human renal glomeruli

A study was undertaken to evaluate some processing variables affecting the immunoelectron microscopic demonstration of immunoglobulins and complement (C3) in human glomeruli. Percutaneous biopsies were performed on 28 patients with various types of glomerulonephritis. Light microscopic, electron microscopic, and immunofluorescence examinations were performed by routine methods. For immunoelectron microscopy, fixation in paraformaldehyde (PA) or periodate-lysine-paraformaldehyde (PLP) was used. With the diffusion technique, using tissue chopper or cryostat sections, human immunoglobulin (Ig)G, IgA, IgM, and C3 were localized in glomeruli with peroxidase-labeled antisera. Using PLP and the tissue chopper sections, good ultrastructure was achieved. The antigens could be demonstrated in intramembranous, subepithelial, subendothelial, or mesangial immune deposits. Penetration of antibodies and quality of peroxidase reaction in the cryostat sections did not differ from that of the tissue chopper sections. Freezing and thawing, however, resulted in inferior morphology. If PA was used, the antigens could not be reliably demonstrated. The results of light microscopy, electron microscopy, and immunofluorescence microscopy were in good agreement with those from the immunoperoxidase procedure. The present study shows that PLP preserves well the antigenicity of human immunoglobulins and C3, resulting in good ultrastructure. PA fixation, on the contrary, caused a loss of antigenicity before an adequate ultrastructure could be achieved.