Histochemical demonstration of some oxidized macromolecules with thiocarbohydrazide (tch) or thiosemicarbazide (TSC) and osmium tetroxide.

Abstract

Thiocarbohydrazide (TCH) and thiosemicarbazide (TSC) have been introduced as osmiophilic reagents for demonstrating aldehyde-containing macromolecules, produced by periodic acid oxidation of tissue sections. Because of the remarkable properties of osmium black, excellent pigment qualities are provided for light microscopy and the advantages of an amorphous, nonvolatile, electron-opaque end product are available for electron microscopy. The present communication describes the method and gives the scope of the histochemical reaction. Thiocarbohydrazide is the more reactive of the two reagents and is capable of demonstrating a wider variety of oxidized macromolecules than does thiosemicarbazide. The periodic acid-thiocarbohydrazide-osmium tetroxide method (PATCO method) was compared with those using thiosemicarbazide (PATO method), periodic acid-Schiff (PAS method), and with the method involving 3-hydroxy-2-naphthoic acid hydrazide (NAH) followed by coupling with fast blue B (PA-NAH-FBB method). All four methods demonstrated certain active aldehydes equally well, such as those produced by periodic acid oxidation of glycogen, mucopolysaccharide of gut and mucoprotein of Descemet's membranes of the cornea. The stain with the PATO method was somewhat weaker than the other three methods for glycoprotein of the cuticular border of gut and of the brush border of kidney tubules, and colloid of the thyroid gland. The PATO method was even less effective in demonstrating mucoprotein of basement membranes in kidney and no reaction for collagen and reticulin was noted. The PATCO method was almost as effective with basement membranes in kidney as the PAS method, but was not equal to the PAS method in staining collagen and reticulin. Furthermore, the aldehyde produced on Feulgen hydrolysis of deoxyribonucleic acid (DNA) of cell nuclei was not demonstrable with either PATO or PATCO methods its contrast to positive results with the PAS and PA-NAH-FBB methods. This suggests that TSC and TCH have less reactive hydrazino groups than NAH. Results with the electron microscope will be published separately.