The presence of phospholipids within the interphase nucleus and in isolated chromatin, previously demonstrated by analytical biochemical methods, has been only rarely documented by cytochemical procedures, especially at the ultrastructural level. By means of a gold-conjugated phospholipase technique, we investigated the fine localization of endogenous phospholipids in the different nuclear domains in rat pancreas and in cell cultures. To reduce possible removal or displacement of phospholipids, different specimen preparation procedures such as cryofixation, cryosectioning, and freeze-fracturing were utilized. Apart from slight differences in efficiency among these methods, phospholipids have been cytochemically identified in the same nuclear domains: the interchromatin granules and fibers and the dense fibrillar component of the nucleolus. These results suggest that the phospholipids are an actual nuclear component, not randomly distributed in the nucleoplasm but mainly localized in the nuclear domains involved in the synthesis, maturation, and transport of ribonucleoproteins.