The choice of a masking agent in the histochemical staining of metals

The masking effects of standard masking agents (aminopolycarboxylic acids, carboxylic acids and phosphates) have been investigated in both test-tube experiments and tissue sections in order to ascertain the factors which must be considered when choosing a masking agent for the histochemical staining of a metal. The masking effects in vitro were determined by spectrophotometry through the complexing of the dye Chrome Azurol S with aluminium, beryllium, and iron at pH 5 and 7. The effects were also examined by staining metal-containing tissue sections in a Chrome Azurol S masking agent system at the same pH values. In many cases, the masking effects observed in sections did not agree with those obtained in the test-tube experiments. This means that the published values of stability constants are not a sufficient guide for choosing a suitable masking agent for the staining of metals. The discrepancy is mainly attributable to the presence of protein in a solid state when metals are stained in sections. Therefore, in the future, consideration should be given to a metal-protein or masking agent-protein interaction using a model compound such as a chelate resin. The polyphosphates are among the most useful masking agents for metal staining in acidic solutions from a practical standpoint.