Histochemical studies of the formazan reaction: III. The reversible oxidation‐reduction of mucosubstance formazans

Abstract

SYNOPSIS In sections of fixed tissue, periodate‐oxidized mucosubstances that have been treated firstly with Phenylhydrazine and secondly at a pH near 9 with a diazonium salt, such as diazotized aniline or tetrazotized 3,3′‐dimethoxybenzidine, are converted into highly‐coloured derivatives which could be formazans, azo‐compounds, tetrazenes, or a mixture of these. That such derivatives are in fact mostly, if not exclusively, formazans has been confirmed by treating these derivatives in situ firstly with N‐bromosuccinimide, to oxidize acid‐labile formazans into acid‐stable, colourless tetrazolium salts; secondly, with a hot ethanolic solution of hydrochloric acid, to destroy azo‐derivatives and tetrazenes and leave tetrazolium salts untouched; and thirdly, with an aqueous solution of ammonium sulphide, to reduce the mucosubstance tetrazolium salts back to the original coloured formazans.