The Feulgen-like osmium—ammine reaction as a tool to investigate chromatin structure in thin sections

• Published in: Journal of ultrastructure research Vol. 80; no. 2; pp. 133 - 147
• Main Authors: Derenzini, Massimo, Viron, Annie, Puvion-dutilleul, Francine
• Format: Journal Article
• Language: English
• Published: United States Elsevier B.V 01.01.1982
• Subjects: Animals; Chickens; Chromatin - ultrastructure; Coloring Agents; Deoxyribonuclease I; Endodeoxyribonucleases; Erythrocytes - ultrastructure; Glutaral; Liver - ultrastructure; Male; Microscopy, Electron; Osmium; Quaternary Ammonium Compounds; Rats; Rats, Inbred Strains; Rosaniline Dyes; Staining and Labeling
• ISSN: 0022-5320; 1878-2345
• DOI: 10.1016/S0022-5320(82)90013-2

The selectivity of the Feulgen-like reaction with the osmium—ammine complex ( Cogliati and Gautier, 1973) and the degree of resolution offered by the staining procedure has been investigated in an ultrastructural study. After HC1 hydrolysis the osmium—ammine/SO 2 stained only the nuclear components known to contain DNA in thin sections of rat liver fixed either with glutaraldehyde or formaldehyde and embedded in Epon. If HCl hydrolysis was omitted, however, osmium—ammine/SO 2 stained both chromatin and cytoplasmic ribosomes in thin sections of glutaraldehyde-fixed tissue. On the contrary, after exclusive osmium—ammine/SO 2 treatment, no cellular structures were stained in formaldehyde-fixed tissue. DNase digestion in frozen sections from formaldehyde-fixed samples also eliminated the Feulgen-like reaction. Chromatin fibers with a thickness of ≈12 nm were the thinner structures visualized by the Feulgen-like reaction in thin sections of glutaraldehyde-fixed samples. No substructural organization could be detected within them. When visualized in thin sections of formaldehyde-fixed tissue, however, the 12-nm fibers showed an inner structure characterized by electron-opaque, ≈3-nm-thick filaments which, irregularly twisted, often delimited electron-translucent areas. Chicken erythrocyte nuclei swollen at low ionic strength were also observed in thin sections from formaldehyde-fixed samples after the Feulgen-like reaction. Chromatin appeared to be characterized by closely associated roundish units, with a diameter of ≈12 nm, composed of electron-opaque DNA filaments, ≈3 nm thick, which encircled electron-translucent areas. It was concluded that formaldehyde fixation is mandatory for the selective staining of DNA in the Feulgen-like reaction with osmium—ammine/SO 2 complex and that this staining procedure, by visualizing DNA structures up to the molecular level, may be considered as a tool to investigate the chromatin structure in thin sections.