Factors affecting lead capture methods for the fine localization of rat lung acid phosphatase.

Abstract

Gomori's lead capture method for acid phosphatase localization was adapted for the electron microscope by Holt & Hicks (1961a). The method gave good results in rat liver, but poor tissue preservation with no reaction product in rat lung, and was, therefore, investigated in order to find the optimum conditions for the ultrastructural localization of rat lung acid phosphatase. The conditions investigated included the use of glutaraldehyde or depolymerized paraformaldehyde as the fixative, with and without dimethylsulphoxide; the effect of freezing the tissue; the pH of the incubation medium; and the use of glycerophosphate, naphthol AS-BI phosphate or α-naphthyl phosphate as substrates. Improved preservation of ultrastructure with increased yield of reaction product was obtained by prefixing lung in glutaraldehyde containing 10% dimethylsulphoxide, freezing the tissue and incubating at pH 5.7 with α-naphthyl phosphate. Tissue preservation was acceptable and dense deposits of reaction product occurred in lysosomal elements of all the alveolar cells and especially in macrophages. Deposits were also found closely associated with the lamellae of the inclusion bodies of Type II cells.