S 450 sem

The 5 kinds of biomineral powders were individually pasted onto the experimental bench and observed using a scanning electron microscope (S-450 SEM, Hitachi, Tokyo, Japan) after gold spraying. The SEM observation of the scaffolds is similar to this. The prepared scaffolds are cut into small pieces and pasted on the test bench. After spraying gold, the scaffolds can be observed by SEM. The SEM images of scaffolds were analyzed by Image J software, and the information regarding fiber diameter and aperture of different scaffolds was counted.

At 120 h of treatment, the metacestodes were fixed and processed for scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Briefly, the metacestodes were fixed in 2.5% glutaraldehyde in PBS (pH 7.2) for 24 h at 4 °C, followed by postfixation in 2% osmium tetraoxide (OsO₄) in 100 mM PBS (pH 7.2) for 2 h at room temperature. For SEM analysis, the samples were washed in distilled water and dehydrated in various concentrations of ethanol (50, 70, 80, 90 and 100%) for 10 min at 4 °C. Subsequently, they were immersed in 2% isoamyl acetate, dried to a critical point and sputter-coated with gold. Finally, they were observed and photographed with a Hitachi S-450 SEM, with an acceleration voltage of 30 kV. For TEM analysis, the samples were washed in distilled water and treated with 1% uranyl acetate for 1 h. They were then rinsed extensively in distilled water, dehydrated in various concentrations of ethanol (50, 70, 80, 90 and 100%) and were embedded in Epon 812 resin. The embedded samples were dried by heat with serial temperatures (40 °C for 48 h and 60 °C for 48 h). Then they were cut into ultra-thin sections with a LKB-Nova ultramicrotome (LKB, Bromma, Sweden), mounted on a copper mesh grid, and stained with 3% uranyl acetate and lead citrate. Afterwards, the samples were observed at an acceleration voltage of 80 kV, in JEOL JEM-1230 TEM.