METHOD FOR ENHANCEMENT OF ELECTRON MICROSCOPIC VISUALIZATION OF EMBEDDED ANTIGEN BY BRIDGING OSMIUM TO URANIUM ANTIBODY WITH THIOCARBOHYDRAZIDE

Abstract

Thiocarbohydrazide (TCH), a bidentate ligand, capable of bridging osmium to osmium in ultrathin sections (OTO method), was shown to possess the additional property of binding osmium to uranium-labeled antibody. On sections in which acrolein-fixed antigen had been localized for electron microscopy by uranium antibody, application of TCH followed by osmication (T-O procedure) greatly enhanced the specific contrast afforded. In addition, the T-O procedure yielded better resolution and better stability to the electron beam than the immunouranium technique employed without this intensification. Even osmium tetroxide-fixed tissue could be used, providing thiosemicarbazide (TSC) was employed to block the later reaction of TCH with tissue-bound osmium and to limit, thereby, the ligation of TCH to the uranium-labeled antibody.