A negative contrast stain for ultra-thin frozen sections

Abstract

Ultra-thin frozen sections are ideal substrates for immunolabelling in high resolution electron microscopy. However, visualization of subcellular structures is inferior to that obtained with corresponding plastic sections. Although negative staining is generally effective and even superior to positive staining, the accumulated stain is often too heavy, obscuring morphology and markers used for immunocytochemical localization of antigens. This paper describes the development of a modified negative contrast staining technique in which a high concentration of uranyl acetate is mixed with methyl cellulose at a low pH. Application of this stain to cryosections of cells and tissue resulted in improved visualization of morphological structures characterized by negative images of membranes and cell organelles. Use of this stain is advantageous for morphological and immunocytochemical studies involving ultra-thin frozen sections.