Resin embedment of organs and postembedment localization of antigens by immunoperoxidase methods

Summary Various procedures for nonpolar and polar resin embedment were applied to mouse and rat livers for the study of postembedment immunolocalization of alpha₁-fetoprotein, albumin and the microsomal enzyme epoxide hydrolase. Fixations with formaldehyde and with formaldehyde-glutaraldehyde mixtures were used for tissue stabilization. Both fixation schedules did not abolish immunoreactivity. Treatment of liver with inert compounds such as polyvinylpyrrolidones or chemical modification of antigens with ethyl acetimidate prior to embedment improved immuno-staining. Either the low-polarity solvent ethanol or the highly polar ethylene glycol could be employed as dehydrating agents. Antigens were readily localized in sections from Epon 812 embedded livers. For this purpose, polymerized resin had to be partially removed. On the other hand, immunoreactivity of antigens was only faint after embedment in an epoxy resin based on diepoxide octane. Also, antigens reacted faintly in sections from livers which were embedded at 0° C in the polar acrylate-methacrylate based Lowicryl K4M resin. The indirect peroxidase labelled antibody method was as specific and sensitive as the PAP technique. Optimal antigen detection was attained with antibodies isolated by affinity chromatography and purified peroxidase conjugates. Apart from purified immunological reagents, the addition of high molarity sodium chloride and bovine serum albumin to the wash solutions enhanced immunohistological specificity.Supported by the Deutsche Forschungsgemeinschaft (Ku 257/3) Bonn, Federal Republic of Germany