Fei tecnai g2 spirit twin 120 kv transmission electron microscope

Cells were pelleted via centrifugation, and the cell pellet was fixed in 2.5% glutaraldehyde in 0.1 M phosphate buffer (pH 7.4) at 4 °C. After 2 h, the pellet was rinsed three times in 0.1 M phosphate buffer and then post-fixed in 1% osmium tetroxide at 4 °C for 1–2 h, followed by three washes in phosphate buffer. The pellet was dehydrated via a series of ethanol solutions and exposed to two changes of propylene oxide. After the dehydration process, the pellet was infiltrated with 2:1, 1:1, and 1:2 mixtures of EMbed 812 resin and propylene oxide for 1 h each time and then finally embedded in 100% EMbed 812 resin. After polymerization for 24–48 h at 60 °C, ultrathin plastic sections (80-nm thick) were cut at room temperature using a Leica EM UC6 ultramicrotome (Leica Microsystems GmbH, Wetzlar, Germany) and collected on 200-mesh carbon-coated grids. The grids were post-stained with 2% uranyl acetate and 1% lead citrate at room temperature for 15 and 5 min, respectively. The Zeiss LEO912AB 120 kV transmission electron microscope (Carl Zeiss, Oberkochen, Germany) and FEI Tecnai G2 Spirit Twin 120 kV transmission electron microscope (FEI Company, Columbia, MD, USA) were used for transmission electron microscopy observations.

Cells were pelleted via centrifugation and fixed in 2.5% glutaraldehyde in 0.1 M phosphate buffer (pH 7.4) at 4°C. After 2 h, the pellets were rinsed three times in 0.1 M phosphate buffer and then postfixed in 1% osmium tetroxide at 4°C for 1–2 h, followed by three washes in phosphate buffer. The pellets were dehydrated through a series of ethanol solutions and exposed to two changes of propylene oxide. After the dehydration process, the pellets were infiltrated with 2 : 1, 1 : 1, and 1 : 2 mixtures of EMbed 812 resin and propylene oxide for 1 h each time. They were finally embedded in 100% EMbed 812 resin. After the resin had been polymerized for 24–48 h at 60°C, ultrathin plastic sections (80 nm thick) were cut at room temperature using a Leica EM UC6 ultramicrotome (Leica Microsystems GmbH, Wetzlar, Germany) and collected on 200-mesh carbon-coated grids. The grids were poststained with 2% uranyl acetate and 1% lead citrate at room temperature for 15 and 5 min, respectively. A Zeiss LEO912AB 120 kV transmission electron microscope (Carl Zeiss, Oberkochen, Germany) and a FEI Tecnai G2 Spirit Twin 120 kV transmission electron microscope (FEI Company, Columbia, MD, USA) were used for transmission electron microscopy observations.