A silver methenamine method for the demonstration of biologic calcification sites

Abstract

Much work has been carried out in our laboratories with the periodic acid-thiocarbohydrazide silver methenamine(PATS) reaction, an improved variation of the periodic acid-Schiff(PAS) or periodic acid-thiocarbohydrazide(TCH)-osmium tetroxide(PATCO) reactions. The PATS stain was used in many of our studies on the vascularization of hydroxylapatite/plaster of Paris implants for bone restoration. It was very useful in demonstrating the Type III collagen or reticulin of the pericapillary reticular network of the newly formed blood vessels. In the course of these studies the improved ability of silver methenamine solutions applied under microwave irradiation to deposit silver at sites of calcification or certain calcium compounds in our specimens was noted. Much more silver was deposited at these sites in femur sections of 5 day old mice after application of 1% silver methenamine soln (pH 9.6) for 1 min at 2450 MHZ and reducing with TCH for 30 seconds (Figs. 1,3,4) than by application of 5% silver nitrate for 30 minutes under bright light and reduction for 2 1/2 minutes with 5% sodium thiosulfate (Fig. 2).