On the cytochemical demonstration of basic proteins in the cell nucleus, including the nucleolus

A combined Feulgen-alkaline fast green method is described for simultaneous demonstration of DNA and basic proteins in the cell nucleus. The method is based on preserving both types of substances in the tissue section and releasing in them reactive groups for the 2 kinds of staining. These conditions are best provided, as proved by staining tests on tissue hydrolysates, if formalin-containing mixtures (SERRA's or LILLIE's fluids) are employed for fixation, and cold 5 N HCl is used for FEULGEN hydrolysis. In this way, a good cytological picture is also achieved. Nuclear euchromatin stains with this method red, while heterochromatin, pycnotic nuclei and sperm heads exhibit a deep violet to blue-violet colour. Prominent nucleoli of metabolically active cells display a distinct blue-green staining thus manifesting their high content of basic proteins. Acetylation test reveals that these proteins are of lysine-rich type. The known negative reaction of the nucleoli with the routine alkaline fast green method according to ALFERT and GESCHWIND must be attributed to an extraction of the nucleolar basic proteins with the hot TCA used in this method. Certain analogy in the cytochemical behaviour between the nucleolous and the chromatin under various conditions of hydrolysis leads to the suggestion that the nucleolar basic proteins demonstrated should be in the form of a ribonucleoprotein complex, probably of the pre-ribosomal material of the nucleolus.