Comparison between light and scanning electron microscopic findings in cells stained by Riu's method.

Difficulties with cell recognition are sometimes encountered in surface structural studies of biomedical specimens with scanning electron microscopy (SEM). If cells are stained and examined first by light microscopy (LM), then the specificity of their surface structures when observed by SEM can be reassured. Although other staining methods have been advocated, Riu's stain, which employs eosin yellow, methylene blue and azure I was tested in this study. A total of 28 specimens, including 10 pleural effusions, 4 ascites, 9 peripheral blood cells and 5 bronchial brushings, were collected for comparison. Free cells were fixed with 2% glutaraldehyde and allowed to form sediment on poly-L-lysine precoated plastic cover slips. They were stained with Riu's A and B solutions and observed with a light microscope. A lattice composed of cut lines made by a surgical knife served as the cell finder. Cells of interest were photographed before they were fixed in OsO4, dehydrated and dried in liquid CO2 at a critical-point temperature. Observation with a scanning electron microscope showed that the particular cells could be found within 10 minutes. Cell loss was minimal. Configuration of the cells was preserved and no distortion was detected in their fine surface structure. This study confirms that Riu's stain is suitable for comparative purposes between LM and SEM.