Increased level of immunogold labeling of epoxy sections by rising the temperature significantly beyond 100°C in the antigen retrieval medium

Abstract The purpose of this study was to compare the level of immunogold labeling of epoxy sections when the sections were subjected to antigen retrieval at different temperatures. Renal swine tissue with glomerular immune complex deposits with reactivity against IgG and C3 was embedded in epoxy resin. Sections from these blocks were exposed to antigen retrieval by heating in citrate solution at temperatures in the range of 25–135°C. Immunogold labeling with anti-IgG and anti-C3 was performed on the heated sections. The level of immunogold labeling increased significantly in the direction of increased heat. Interestingly, the level of immunogold labeling was significantly higher when exposed to heating in the autoclave (121 and 135°C) than at temperatures just below the normal boiling point. Sections stained with anti-C3 turned from almost negative labeling when heated at 95°C to strong positive labeling when heated at 135°C (11 times increased). The intensity of the immunogold labeling with anti-IgG increased almost three times when raising the temperature in the retrieval medium from 95 to 135°C. The practical significance of these results is that antigen retrieval of epoxy sections should be performed by heating in aqueous solutions at 135°C or higher to obtain maximum immunolabeling.