Improved Method of Embedding with Epoxy Resin 'Quetol 651' for Both Light and Electron Microscopic Observation of Identical Sites in Semi-thin Sections

In order examine semi-thin sections appmximately 0.2μm thick for light microscopy with an accelerating potential of 100 kV by a conventional electron microscope, Quetol 651 as a low viscosity water-miscible epoxy resin was suitable for an embedding medium. Semi-thin sections approximately 0.2μm thick embedded in Quetol 651 could be examined with an ordinary electron microscope operating at 100 kV. Tissues were fixed in 2.5% glutaraldehyde with buffered cacodylate at pH 7.4for 3 hr, postfixed in 2% osmium tetroxide with buffered cacodylate at pH 7.4 for 2 hr and stained with en bloc by 3% aqueous solution of uranyl acetate for 2 hr at 37°C. After dehydration in graded alcohol, they were embedded in the Quetol 651mixture. Semi-thin sections approximately 0.2μm thick were cut with glass knives on a conventional ultramicrotome. Semi-thin sections on grids were stained with the Giemsa method and methylene blue-azure II-basic fuchsin. Staining without removal of the embedding matrix was sharp and brilliant. These stained sections were observed under a light microscope. For electron microscopy, they were doubly stained with uranyl acetate and lead salt. Identical sites on such sections could be distinctly examined with an accelerating potential of 100 kV at low magnification (250-1,500 times) using LEM-2000, a combined light and electron microscope. Thus, photomicrographs electron micrographs of identical sites on tissue samples could be compared exactly.