SCANNING ELECTRON-MICROSCOPY OF HEPATIC ULTRASTRUCTURE - SECONDARY, BACKSCATTERED, AND TRANSMITTED ELECTRON IMAGING

  • 1 January 1976
    • journal article
    • research article
    • Vol. 35  (4) , 369-376
Abstract
Several methods of tissue preparation and different modes of operation of the scanning electron microscope [SEM] were used to study the ultrastructure of rat liver. Rat livers were perfusion fixed with buffered 2% paraformaldehyde or a mixture of 1.5% paraformaldehyde and 1% glutaraldehyde. Tissue blocks were postfixed in buffered 2% osmium tetroxide followed sequentially by the ligand-mediated osmium binding [LMOB] technique, dehydration and cryofracture in ethanol, and critical point drying. They were then examined without metal coating in the SEM operated in the secondary electron [SE] and backscattered electron [BSE] modes. Fifty-micrometer sections were cut with a tissue sectioner, stained with lead citrate, postfixed with osmium, dehydrated, critical point dried and examined in the SE and BSE modes. Frozen sections (0.25-0.75 .mu.m/thick) were cut by the Tokuyasu method and their transmission SEM images were examined either with a transmission SEM detector or with a conversion stub using the SE detector. SE images of the liver prepared by LMOB and subsequent cryofracture revealed such intracellular structures as cisternae of the endoplasmic reticulum, lysosomes, mitochondria, lipid droplets, nucleolus and nuclear chromatin, as well as the usual surface morphology. Lipocytes in the perisinusoidal space were readily identified. BSE images revealed some subsurface structures but had much less resolution than that of SE images. Unembedded frozen sections had little drying artifact and were virtually free of freezing damage. The transmission SEM image revealed those organelles visualized by the SE mode in the LMOB-treated tissue.

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