Veleta ccd camera

A series of 50-μm-thick sections−250 μm apart from each other—were collected from the cortex (from Bregma, ~-2.9; lateral, ~±0.25; depth, ~-0.6 mm) of mice. After washing in PBS, the sections were treated with 0.5% osmium-tetroxide for 20 min, dehydrated, and embedded in epoxy resin. During dehydration, the sections were treated with 1% uranyl acetate. After polymerization, we prepared 70-nm-thick sections (Leica EM UC6, Wetzlar, Germany) of the outer two-thirds of the cortex, picked them up on formvar-coated single-slot copper grids, and examined them using a JEOL-1200EX electron microscope (EM) and a Soft Imaging System Veleta CCD camera (EMSIS, Münster, Germany). The synaptic area was measured from serial sections of three-dimensional (3D) reconstructed synapses. We only included synapses that were cut perpendicularly to the sectioning plane.

A series of 50 μm thick sections 250 μm apart from each other were collected from the parietal cortices of the animals. After washing in PBS, sections were treated with 0.5% osmium-tetroxide for 20 min, dehydrated and embedded in epoxy resins. During dehydration, sections were treated with 1% uranyl acetate. After polymerization, we prepared 70 nm thick sections (Leica EM UC7) from the parietal cortex, picked them up on formvar-coated single-slot copper grids and examined them using a JEOL-1200EX electron microscope (EM) and a Soft Imaging System Veleta CCD camera (EMSIS, Münster, Germany). For measurements of images of electron microscopy, we used Image J software.

PERV-F producing 293 T cells were fixed with 2.5% glutaraldehyde in 50 mM 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid (HEPES), pH 7.2. The cells were harvested by scraping, pelleted at 2.000 × g for 5 min at 4 °C, and washed twice with HEPES. After washing, the cells were block-embedded by mixing equal amounts of pelleted cells and low-melting-agarose (3%). Agarose-embedded cells were cut into small pieces (< 1 mm), and postfixed with osmium tetroxide (1% in double distilled H₂O for 1 h), tannic acid (0.1% in 50 mM HEPES for 30 min), and uranyl acetate (2% in ddH₂O for 2 h). The agarose-embedded cells were dehydrated in a graduated ethanol series and finally embedded in Epon resin. Thin sections (60–70 nm) were cut with a Leica UC7 ultramicrotome, using a diamond knife (45°, Diatome, Switzerland), mounted on naked 300 mesh copper grids, and counterstained with uranyl acetate (2% in ddH₂O for 20 min), followed by lead citrate (Reynolds’ solution for 3 min). Ultrathin sections were stabilized with a thin layer of carbon evaporation and examined using a JEM-2100 transmission electron microscope (JEOL) at 200 kV. Images were recorded using a Veleta CCD camera (EMSIS) and 2048 × 2048 pixel.