An established routine method for differential staining of epoxy-embedded tissue sections.

A method is described for staining semithin sections of epoxy-embedded cells and tissues. PAS-positive structures are specifically demonstrated by a red staining providing a vivid contrast to the other tissue components differentially stained in shades of blue, using methylene blue/azure II. Typical staining conditions for 1 to 2 micron thick Epon sections include 30 min oxidation with 5% periodic acid at 50 degrees C, 30 min incubation with Shiff's reagent at 50 degrees C, 20 min counter-staining with 0.5% methylene blue/0.5% azure II in 0.5% aqueous borax solution at room temperature. For 10 years, this method has provided excellent differential staining with a variety of tissues. Stained sections showed no signs of fading during this period of time. Therefore, this procedure is recommended as a simple method of staining semithin sections both for tissue orientation in electron microscopy and for brillant representation of cells and tissues, required for microphotography in color or black-and-white.