X-ray microanalysis of non-aldehyde-fixed glycogen contrast stained with OsVIIIO4, OsVIIIFeIII, or OsVIFeII complex in vitro

Summary Fibrin-enrobed, commercially produced glycogen was treated, without prior glutaraldehyde fixation, to a form of post-fixation with solutions of Os^VIIIO₄ or with a mixture of either Os^VIIIO₄ plus K₃Fe^III(CN)₆ or K₂Os^VIO₄ plus K₄Fe^II(CN)₆.Only the last mixture gave constrast staining of the glycogen in unstained ultrathin sections. The first mixture rendered the glycogen just barely visible but the glycogen contrast was increased by lead staining. The glycogen treated with the Os^VIIIO₄ solution was not contrast stained and was just observable after lead staining.Qualitative X-ray microanalysis of the glycogen in the ultrathin sections confirmed the presence of osmium and iron in the glycogen treated with both mixtures. The glycogen treated with Os^VIIIO₄ alone was difficult to analyse.Quantitative X-ray microanalysis showed that, in the glycogen treated with the Os^VIIIO₄ mixture plus K₃Fe^III(CN)₆, the mean atomic osmium to iron ratio was 15. In the glycogen treated with K₂Os^VIO₄ plus K₄Fe^II(CN)₆ this ratio was 117. However, the mean net osmium intensity in the latter case was 15 times higher than in the former case and for the iron even 40 times higher.The Unit for Analytical Electron Microscopy was established by collaboration between the Erasmus University of Rotterdam (W. C. de Bruijn), the University of Leiden and the Organization for Health Research TNO. The analytical microscope was purchased with funds from the Netherlands Organization for Pure Scientific Research (ZWO).