A Silver-Schiff Reaction for Cytological Studies: Light Microscope Observations

A method is presented in which a complex metal-ion (ammoniacal silver) is substituted for leuco-basic fuchsin in the well-known Schiff reaction for aldehydes. The reagent is reduced to metallic silver by tissue aldehydes and submicroscopic particles of the metal are deposited at the reactive sites. When the reagent is used in the Feulgen reaction chromosomes may be ‘stained’ in a specific manner to show the localization of deoxyribonucleic acid. For such a purpose, fixation should be carried out with acetic acid and alcohol mixtures rather than with solutions containing formalin or dichromate; these latter substances preserve more of the tissue lipids and thus complicate the issue by producing cytoplasmic staining (plasmal reaction).

A Critique of the Plasmal Reaction, with Remarks on Recently Proposed Techniques

1. The acetalphosphatides which are the precursors of the aldehydes (‘plasmal’) responsible for Feulgen's plasmal reaction are extremely labile compounds which hydrolyse very rapidly in acid media, and are destroyed more or less rapidly during fixation. The liberation of aldehydes from them is catalysed by mercuric chloride. 2. Ordinary unsaturated lipoids, if exposed to air, become capable of producing a colour with Schiff's reagent, which, unless a control section is used, can be misinterpreted as a positive result for Feulgen's plasmal reaction. Mercuric chloride has little or no influence over the oxidative rancidity of unsaturated lipoids. 3. Any technique prescribed for showing the plasmal reaction must avoid acid media and prolonged fixation and handling of the tissues. A control section is a necessity.

On the Significance of the Plasmal Reaction

1. Two different reactions have been confused under the name ‘plasmal’. One, the true plasmal reaction, is due, as its discoverer Feulgen showed, to the liberation of higher aliphatic aldehydes from acetalphosphatides, the reaction taking place very rapidly under the influence of mercuric chloride, the products being coloured deeply with Schiff's reagent. The other reaction is due to the oxidation, by atmospheric oxygen, of double bonds in unsaturated fatty acid radicles. It is affected only slightly, if at all, by mercuric chloride, and increases with exposure of sections or pieces of tissue to atmospheric oxygen. 2. There is no completely satisfactory method for the histochemical demonstration of plasmal. Sections of fresh tissues are not usually suitable for cytological work, and fixation causes a reduction in intensity of the reaction. Also, the permissible fixatives are not all of the highest quality. Short fixation in a formaldehyde fixative (e.g. formal-calcium) followed by careful washing is probably the best, but the results should be compared with those obtained by the direct reaction of small pieces of fresh tissue. A suitable technique is described.