The rats were euthanized by intraperitoneal injection of 1% pentobarbital (800 mg/kg), and 1 mm3 of substantia nigra tissues was quickly removed and fixed with 2.5% glutaraldehyde at 4°C for 2 h. After fully washing with PBS, the samples were fixed with 1% Russian acid. After fully washing with PBS, the samples were dehydrated with 30, 50, 70, 90, and 100% ethanol for 15 min, and propylene oxide was added to replace the ethanol for 30 min. Then, the samples were embedded with Epon 812 epoxy resin (Ted Pella, CA, USA), placed in the 1:1 mixture of Epon 812 epoxy resin (formula: Epon 812: DDSA: NMA: DMP30 = 27.5:4:20:0.75) and propylene oxide for 2 h, and in the 2:1 mixture for 1 h, then soaked in pure epoxy resin Epon 812 for 2 h, and placed into a 60°C oven for 12 h. The embedded tissues were prepared into 50- to 70-nm-thick ultrathin sections using an ultrathin slicer (Leica EM UC7, Leica, Germany). With 100-mesh copper mesh as the carrier mesh, the sections were stained with uranyl acetate and lead citrate and observed under a TEM (HITACHI H-7650, HITACHI, Japan) to observe the ultrastructural changes in neuronal mitochondria in substantia nigra of rats.
Epon 812 epoxy resin
Manufactured by Ted Pella
Sourced in United States
Epon 812 is an epoxy resin designed for embedding and embedding samples for electron microscopy. It is a low viscosity, low shrinkage resin that provides good sectioning characteristics and preservation of ultrastructural detail.
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Lab products found in correlation
2 protocols using epon 812 epoxy resin
1
Ultrastructural Analysis of Mitochrondria in Substantia Nigra
2
Ultrastructural Analysis of Sperm Exposed to SIN-1
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The analysis of sperm ultrastructure was performed by transmission electron microscopy (TEM) using a Zeiss EM 900 transmission electron microscope (Zeiss, Oberkochen, Germany). For the experiments, sperm aliquots (2 × 106 spermatozoa per ml) from three different donors previously exposed to SIN-1 and controls were fixed for 2 h at 0°C–4°C with fixative solution consisting of 4% paraformaldehyde (w/v), 4% glutaraldehyde (w/v), and 20% picric acid (v/v) saturated in PBS. Fixed sperm were washed twice with PBS for 10 min at 600 g. Then, the pellet containing the spermatozoa was postfixed by adding 30 ml of 1% osmium (VIII) oxide (OsO4; w/v) and incubated overnight at 4°C. Osmified samples were dehydrated in ethanol-acetone up to absolute acetone and embedded in Epon 812 epoxy resin (Ted Pella Inc., Redding, CA, USA). Ultrathin sections were obtained by Ultracut R microtome (Leica Biosystems, Vienna, Austria) and stained with classical uranyl acetate and lead citrate TEM stain. The ultrathin sections were examined with a TEM at 80 kV. The results were presented as percentage of alterations on sperm ultrastructure. Representative images of each condition were also obtained.
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