Use of distilled water as a rinsing solution for intracellular observation by scanning electron microscopy

A new specimen preparation to reveal intracellular structures is introduced and its application to some tissues is demonstrated. By this method, intracellular structures are easily observed by SEM. Moreover, some parts of the cells and tissues are cleaved and dissociated, and interesting structures are revealed. Tissues are fixed with 1% osmium tetroxide, rinsed with distilled water and freeze cracked with 50% DMSO. The cracked specimens are rinsed with distilled water to remove cytoplasmic matrix. The author calls this method "Osmium-DMSO-distilled Water method (O-D-W method)". When prepared by this method, intracellular structures such as mitochondria and endoplasmic reticulum are easily demonstrated three-dimensionally by SEM. Sometimes, the nucleus is removed during the specimen preparation procedure and endoplasmic reticulum around the nucleus is seen three-dimensionally. When cells and tissues are cleaved and dissociated, lateral surface of the intestinal epithelium and the reverse side of the microvillous border is also observed. Peritoneal cells lining the liver is also detached from the basement membrane and the surface of the basement membrane is exposed.