Observation on backscattered electron image (BEI) of a scanning electron microscope (SEM) in semi-thin sections prepared for light microscopy.

In order to examine semi-thin section for light microscopy with the backscattered electron mode (BE mode), identical sites in tissue sections were comparatively observed with both light microscopy and BE mode. Tissue blocks (ca. 3 X 3 X 1 mm) were fixed in glutaraldehyde or combined formaldehyde-glutaraldehyde solution. After dehydration in alcohol, they were embedded in Kushida's GMA-Quetol 523. 1.0 micron sections on glass slides coated with indium oxide were stained with hematoxylin-eosin or toluidine blue or by the Giemsa method, and then treated with osmium tetroxide vapor or aqueous KMnO4 solution or uranyl acetate-lead citrate solution. The identical places of such sections could be examined with the accelerating potential of 6 kV and the probe current of 8 X 10(-10) A using a JSM-35C SEM with BEIS BE detector. Photographs were taken with the 2500-line resolution cathode ray tube and the time of exposure was 100 sec. The sections were placed at a distance of 5mm from the BE detector. BE images from osmium tetroxide vapor staining showed a distinctly improved contrast especially when the sections were previously stained with hematoxylin and eosin. The cellular structure was clearly demonstrated under the electron microscope in the BE mode. Identical sites in tissue samples could be compared exactly with both light and electron micrographs.