Type of presentation: Poster

LS-2-P-2321 Staining in non-isotope metals to cell structure in serial sections from animal tissues

Muranaka Y.1, Nishida T.1, Park S. M.2, Park P.3
1Research Center for Ultra-High Voltage Electron Microscopy, Osaka University, Japan, 2Research Center for Environmental Genomics, Kobe University, Japan, 3Emeritus Professor of Kobe University, Japan
muranaka@uhvem.osaka-u.ac.jp

Uranium has been used widely as the pre-stain of electron double stain for electron microscopy. However, the use and purchase have become more difficult in recent years. Therefore, the study on developing alternative uranium is underway. Some non-isotopic heavy metals such as hafnium chloride (HF), samarium chloride (SC), platinum blue (PB) were recommended in other studies as alternative stains of uranyl compounds to TEM sections from aldehyde-and osmium-fixed biological tissues. The cell contrast in tissues stained with the metals and lead was compared in the nearly same spatiality sites of serial sections with that in tissues with uranyl acetate (UA) and lead (Pb). The stains used were in the six followings: 2% UA aqueous solution and 50% EtOH, 4% HF in 50% and 100% EtOH, 4% SC solution, Pb solution while the test tissues were liver, kidney, small intestine, nerve, skeletal muscle of mice. Seven serial sections with 60 nm in thickness were cut from each of the blocks. Each of sever sections were stained with different staining methods used here. The contrast of cell images due to different stains was examined with penetration rates of stains in sections. The staining mechanisms of stained in nucleic acid and proteins will be discussed from the result obtained. In addition, the non-isotope staining solution are explored to replace the uranium in field of the ultra-high voltage electron microscope such as the method of three-dimensional reconstruction. Information of penetration of the stain and the contrast of the image is very useful.

Fig. 1: Hepatocytes TEM image stained with 2% uranium acetate in aqueous solution. Ultrathin sections of the three following the ultra thin section in Figure 1, is used 2, 3 and 4.

Fig. 2: Hepatocytes TEM image stained with 4% hafnium chloride in 50% alcohol solution. Staining of the cell substrate is comparable to uranium acetate.

Fig. 3: Hepatocytes were stained with TI blue stock solution was diluted 5-fold with 50% alcohol. in comparison with other stains, Ti blue is difficult to obtain the contrast such as staining of chromatin is low.

Fig. 4: Hepatocytes were stained with 4% aqueous solution of chloride Samaniumu. Staining closest to uranium is obtained. Necessary to study the permeability of the resin, the staining of the different organizations.