Methods in ultrastructural cytochemistry of the cell nucleus

Abstract

The electron microscopical study of the cell nucleus as observed in thin sections requires the use of cytochemical methods because of the intricate pattern of the nuclear components. The in situ techniques based on electron staining and enzymatic digestion are reviewed, excluding autoradiography, cytoenzymology and immunocytochemistry. A tentative classification has been adopted according to the chemical nature of the revealed component. Thus, the staining procedures for the nucleoproteins in general, for both nucleic acids, for the proteins, and finally for the deoxyribonucleoproteins and DNA are considered separately. 1--Stains for the nucleoproteins include simple reagents such as the uranyl and lead salts which are largely used in electron microscopy but are of limited specificity. 2--A variety of methods, some of them specific, is available for the simultaneous visualization of DNA and RNA which is based on common properties: basophilia, ability to bind diaminoacridines, presence of hydroxyl groups. However, due to the recent development of specific and preferential methods for each nucleic acid, we feel that among the older methods, only rapid and simple procedures for the detection of both nucleic acids remain of interest. 3--Proteins being ubiquitous, the useful techniques must reveal subsets within the total nuclear proteins. Apart from some endogeneous enzymes, basic proteins -- practically histones -- so far represent the only group for the detection of which reliable methods exist. 4--Several techniques developed recently are available for the specific detection of DNA. In favourable cases, methods derived from the Feulgen reaction allow its visualization at a molecular level. In addition, standard procedures for the preparation of mammalian cells and tissues are described. Each staining method is at least briefly discussed, but emphasis has been placed on a small number of techniques described in detail. They comprise the EDTA regressive stain for the ribonucleoproteins, several reactions of the basic proteins and the Feulgen-like osmium ammine reaction for DNA.