The contribution of osmium tetroxide to the image quality and detectability of elements in cells studies by electron spectroscopic imaging

Abstract

Osmium tetroxide has been recognized as an excellent fixative which also contributes to the quality of TEM micrographs (1). In the preparation of biological materials for electron spectroscopic imaging (ESI) of 30 run sections with the Zeiss CEM902 electron microscope, OsO4 is usually used in fixation, but poststaining with heavy metals is not needed. However, the extent of the contribution of OsO4, to the images obtained had not been defined clearly, nor was it clear the extent to which the presence of OsO, might obscure detection of elements by ESI or EELS. To establish optimal liquid fixation for the preservation of soluble ions we monitored the loss of Fe ions during preparation. To determine the role of Os4O, in ESI imaging, hamster alveolar macrophages (AMS) were incubated with 0.ImM FeCLo for 15 minutes, then fixed with 2.5% glutaraldehyde in 0.IM K phosphate buffer in 0.01% CaCLo, then grouped into 6 samples varying the times and percentages of OsO, . One sample was not exposed to any OsO4.