Staining Reactions of Collagen in ‘Epon’ Sections

Abstract

IT was recently reported¹ that collagen fibrils appeared tubular in electron micrographs of ‘Epon’ sections stained with Karnovsky's² alkaline lead hydroxide solution, and it was speculated that mucopolysaccharide might be present within the fibrils. The colloidal iron method for demonstrating acid mucopolysaccharides^3,4 has been adapted for the electron microscope^5,6, and although it confirms that acid mucopolysaccharide is present in the ground-substance coating the fibril (Fig. 1) it provides no evidence for the presence of mucopolysaccharides within the fibril. It seems that two factors, which may be interrelated, contribute to the tubular appearance of the fibrils: (1) ‘Epon’ penetrates collagen fibrils poorly and sometimes not at all; (2) the lead hydroxide solution tends to have a stronger affinity for ground-substance than for collagen fibrils (Figs. 1 and 2). Nevertheless, longitudinal sections still show the characteristic banding of collagen (Fig. 3), which means that the ground-substance coating each fibril is acting like a stained replica of the fibril's surface. This affinity of lead for ground-substance is found after fixation in osmium-sucrose solution, formalin, glutar-aldehyde and Luft's solution. Oddly enough, it can be influenced by the process of section-cutting, and wherever the ‘Epon’ has been compressed during cutting there are occasional areas in which the image is reversed, with dark-stained collagen fibres and pale ground-substance.