Ultrastructural visualization of elastic fibres with a tannate-metal salt method

Abstract

A modification of the tannic acid-metal salt method was applied as an ultrastructural stain for elastin. Thin sections of glutaraldehyde-fixed, embedded rat aorta and rabbit elastic cartilage, with and without osmication, were examined. Raising the pH of the tannic acid solution from 2.7 to 9.0 progressively increased the electron-density of elastic fibres and collagen fibrils in osmicated and unosmicated specimens. The maximum tannic acid staining of elastic fibres was observed in the pH range 7.0-9.0. Collagen staining, although less intense than that of elastic fibres, was also greatest in this pH range. Elastic fibres in osmicated specimens demonstrated the strongest tannic acid staining with a minimal increase in density of collagen and cell nuclei when compared to the unosmicated specimens. Sequential treatments of osmicated specimens with tannic acid pH 7.0-9.0, and uranyl acetate, pH 4.1, enhanced the density of the elastin intensely, increased collagen staining moderately, but hardly increased the density of nuclei and microfibrils. In elastase-digested osmicated specimens, all tannic acid (pH 7.0)-uranyl acetate-reactive elastin was selectively removed. These results demonstrate that all the neutral and alkaline tannic acid-uranyl acetate methods can be used as a postembedment stain for elastin specimens fixed in glutaraldehyde and osmium tetroxide.

Three-dimensional regular arrangement of the annular ligament of the rat stapediovestibular joint

“…Furthermore, we have confirmed that the electronlucent central amorphous substance has a high electron density by the tannate-metal salt method (Kageyama et al, 1985). These results clearly indicate that the ligament fibers of the SVJ are mainly composed of mature elastic fibers.…”

“…Ultrathin sections were cut with a diamond knife, mounted on gold grids, and contrasted with a saturated uranyl acetate aqueous solution for 10 min (Stempak and Ward, 1964) and Reynolds' lead citrate solution for 40 s (Reynolds, 1963). For visualization of elastic fibers, the modified tannate-metal salt method was used (Kageyama et al, 1985). Ultrathin sections mounted on gold grids were treated for 5 min with a filtered, freshly prepared tannic acid solution, containing 5 g of tannic acid in 95 ml of distilled water (pH 7.0, adjusted with NaOH), rinsed in three successive baths of distilled water for 1 min and immediately followed by uranyl acetate and lead citrate staining as described above.…”

Quantitative analysis of chondroitin sulphate retention by tannic acid during preparation of specimens for electron microscopy

“…Bar = 1 ~m. Kageyama et al, 1985), this is the first study to show its ability to enhance binding of chondroitin sulphate to collagen in a synthetic model system. TA was shown here to enhance preservation of the original mixture of macromolecules in situ in the synthetic model, and we hypothesize that this same effect might have to do with the structural reinforcement bestowed by TA on the surfaces of resected joint-apposing tissues, reported previously (Levanon & Stein, 1991, 1992, 1993.…”

Nitrogen dioxide induces apoptosis and proliferation but not emphysema in rat lungs

“…23 For quantification of the volume fractions of the components of the alveolar wall-that is, alveolar epithelium, capillary endothelium, and interstitial tissue including elastin and collagen deposition-point counting was performed on each upper left corner of the grid. A transparent sheet with 140 equidistant test points was superimposed onto a television monitor, which was connected to a Zeiss EM 900 transmission electron microscope (Carl Zeiss, Oberkochen, Germany), and used for counting all points hitting the respective component relative to the total number of hits on the alveolar wall at a final magnification of 297 (13)*À**1ÀÀ 18.6 (2.7)À**1ÀÀ 10 ppm NO 2 3 6 189 (9)À`ô 5.8 (3.6)*À`ÀÀô 10 ppm NO 2 7 6 183 (5)*À`ÀÀô 7 (2.8)*À`ÀÀô 10 ppm NO 2 21 6 205 (11)À`1ô 9.4 (1.8)*À`**1 10 ppm NO 2 /room air 21 6 201 (8)À`ô 9.2 (1.9)*À`**1 + 28 280 (11)*À**1ÀÀ 19.8 (2.1)*À**1…”