Es500w ccd camera

Cells were grown in SD medium. Once they reached OD₆₀₀ ~ 0.5, 5 mL of a fixative solution (6% paraformaldehyde, 4% glutaraldehyde and Cacodylate buffer 0.2 M (pH = 7.4)) was added to 5 mL of media. The samples were gently rotated for 40 minutes at 30 °C, then centrifuged and washed twice in Cacodylate buffer and incubated again for an hour in the fixative solution. After washing with Cacodylate buffer and centrifugation, samples were embedded in 10% gelatin in water, fixed overnight with the fixative solution, washed in Cacodylate buffer, and then incubated overnight in 2.3 M sucrose and rapidly frozen in liquid nitrogen. Frozen ultrathin (70–90 nm) sections were cut with a diamond knife (Diatome AG, Biel, Switzerland) at –120 °C on an EM UC6 ultramicrotome (Leica Microsystems, Vienna, Austria). The sections were collected on 200-mesh Formvar-coated nickel grids. Contrasting and embedding were performed as previously described [46 (link)]. The embedded sections were observed in a Tecnai T12 electron microscope (FEI, Eindhoven, The Netherlands) operating at 120 kV. Images were recorded using an Erlangshen ES500W CCD camera (GATAN) or an Eagle 2 k × 2 k CCD camera (FEI). The quantification of mitochondrial area and cristae was performed using imageJ [47 ].

Sciatic nerves fixed in 2% glutaraldehyde/1% paraformaldehyde and post-fixed in 1% osmium tetroxide were Epon-embedded using routine procedures, and ultrathin sections were cut. Semi-thin sections (0.5 μM) were used for toluidine blue staining, and ultrathin sections were used for electron microscopy. The ultrathin sections were mounted on formvar-coated copper grids and contrasted with uranyl acetate and lead citrate. The samples were viewed in a Jeol JEM-2100 electron microscope at an acceleration voltage of 120 kV and a magnification range of 1,500× to 4,000×. Electron micrographs were recorded on a Gatan Erlangshen ES500W CCD camera.