Quarterly Journal of Microscopical Science, Vol s3-96, 84-102, Copyright © 1955 by Company of Biologists

Experiments on the Mechanism of Silver Staining

Part I. Impregnation

¹ Department of Zoology, University of Bristol

In the impregnating stage of silver staining, there are essentially two different reactions taking place between the silver ions and the sections of fixed tissue. A substantial part of the silver is combined with the tissue in the unreduced form, and a smaller part is reduced to produce silver nuclei which form centres on which the developing gents can deposit additional silver derived from the combined (reducible) silver fraction.

A study of the effect of_pH and time of impregnation, as well as of the specific blocking of chemically active protein end-groups upon the reducible silver fraction, suggests that this is chiefly combined with histidine and that it is not limited to the sites of the final differential staining. Except for the cell nuclei, the reducible silver shows a uniform distribution throughout the section; the amount taken up is in equilibrium with the silver ion concentration in the impregnating bath.

The fraction present as silver nuclei increases progressively with time, with increasing pH, and with the temperature ofthe impregnating bath. Silver ions appear to be converted to silver nuclei by a process of physical reduction, and once formed, the nuclei are quite stable.

Deposition of silver nuclei, necessary for differentiation of the stain, only occurs after impregnation between pH 7.5 and 9.0. In addition, it is only within this range that a sufficient amount of reducible silver is taken up to produce deep staining after development.