Transmission electron microscopy was performed as described previously [56 (link)]. Briefly, 2.5% glutaraldehyde-fixed cells were post-fixed with 1% osmic acid (Sigma-Aldrich, O5500) followed by acetone dehydration. Dehydrated pellets were embedded in araldite CY212 for sectioning, followed by staining with alcoholic uranyl acetate (Polysciences, Inc., 6159–44-0) and alkaline lead citrate (Sigma-Aldrich, 15326). Autophagic vacuoles of the ultrathin sections were examined under a transmission electron microscope (JEOL Ltd, Tokyo, Japan, JEM-1230).
O5500
Manufactured by Merck Group
Sourced in Germany
The O5500 is a laboratory equipment product from Merck Group. It is designed for general laboratory use, providing core functionality for various applications. The details of its intended use and specific applications are not provided in this factual description.
Automatically generated - may contain errors
Lab products found in correlation
G5882, by Merck Group (2 mentions)
Toluidine blue, by Merck Group (2 mentions)
Jem 1230, by JEOL (1 mentions)
Alkaline lead citrate, by Merck Group (1 mentions)
Jem 1200ex transmission electron microscope, by JEOL (1 mentions)
G1102, by Wuhan Servicebio Technology (1 mentions)
Ht7700, by Hitachi (1 mentions)
Agr1260a, by Agar Scientific (1 mentions)
Morgagni 268d, by Thermo Fisher Scientific (1 mentions)
G6257, by Merck Group (1 mentions)
Potassium ferrocyanide, by Merck Group (1 mentions)
C0250, by Merck Group (1 mentions)
Sodium borate, by ITW Reagents (1 mentions)
9 protocols using o5500
1
Transmission Electron Microscopy of Autophagic Vacuoles
2
Transmission Electron Microscopy Protocol
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Cells were fixed with glutaraldehyde (2.5%), immobilized in osmic acid (1%, O5500, Sigma‒Aldrich), dehydrated with acetone, embedded in Araldite CY 212 (E009, TAAB), and stained with alcoholic uranyl acetate and alkaline lead citrate. The sections were observed under a JEM 1200EX transmission electron microscope (JEOL Ltd, Tokyo, Japan).
3
Ultrastructural Analysis of Cochlear Hair Cells
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The cochleae were fixed in 2.5% glutaraldehyde fixative (Servicebio, G1102) overnight, decalcified using 10% EDTA decalcification solution (Servicebio, G1105) at room temperature for 1 week, and fixed in 1% osmic acid (Sigma‐Aldrich, O5500) at room temperature for 2 h. After gradient dehydration, the cochleae were embedded in epoxy resin parallel to the cochlear axis. Semi‐thin sections with a thickness of 2.0 μm were taken, and 1% toluidine blue staining was performed to detect the intact cochlear axis. Ultrathin sections were prepared using an ultramicrotome (Leica, UC7). These sections were double‐stained with 3% uranyl acetate and lead citrate for 15 min. A transmission electron microscope (JEM‐1200EX, Japan) was then used to observe the morphology and mitochondrial ultrastructural characteristics of these cochlear hair cells.
4
Ultrastructural Analysis of Tumor Mitochondria
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Briefly, tumors were cut into pieces of about 1 mm3, and cells were grown on coverslips. Samples were fixed with 2.5% glutaraldehyde (Sigma-Aldrich, G5882) for 2 h at room temperature. After wash, samples were post-fixed with 1% osmium tetroxide (Sigma-Aldrich, O5500) for another 1.5 h, and dehydrated with an ethanol series. Samples were infiltrated, embedded in Epon Resin, and polymerized at 60 °C for 12 h. Ultrathin sections of 60 nm were prepared, stained with uranyl acetate and lead citrate. Samples were observed under a Hitachi HT7700 transmission electron microscope. Mitochondrial length was analyzed by an Image-Pro Plus 6.0 software. For quantification of MAM, we normalized the MAM region to total mitochondrial perimeter following a published method62 (link).
5
Electron Microscopy Tissue Preparation
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Glutaraldehyde-fixed tissue specimens were rinsed in cold cacodylate buffer (20840; Sigma-Aldrich), postfixed in 1% osmium tetroxide (O5500, Sigma-Aldrich), dehydrated in graded ethanol and propylene oxide (82320, Sigma-Aldrich), and processed for embedding in EPON (45345; Sigma-Aldrich). Thin sections were mounted on copper grids and stained with uranyl acetate (AGR1260A; Agar Scientific Ltd., Essex, UK) and lead citrate (AGR1210; Agar Scientific Ltd.) and further examined by electron microscope (Morgagni 268D; FEI, Hillsboro, OR, USA).
6
Ultrastructural Tissue Preparation Protocol
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EC sections were postfixed for 24 h in a solution containing 2% paraformaldehyde, 2.5% glutaraldehyde (TAAB, G002, UK) and 0.003% CaCl2 (Sigma, C-2661-500G, Germany) in sodium cacodylate (Sigma, C0250-500G, Germany) buffer (0.1 M). These sections were washed in sodium cacodylate buffer (0.1 M) and treated with 1% OsO4 (Sigma, O5500, Germany), 0.1% potassium ferrocyanide (Probus, 23 345, Spain) and 0.003% CaCl2 in sodium cacodylate buffer (0.1 M) for 1 h at room temperature. After washing in PB, sections were stained with 2% uranyl acetate (EMS, 8473, USA), and then dehydrated and flat-embedded in Araldite (TAAB, E021, UK) for 48 h at 60°C (DeFelipe and Fairén 1993 (link)). Embedded sections were glued onto a blank Araldite block and trimmed. Semithin sections (1–2 μm thick) were obtained from the surface of the block and stained with 1% toluidine blue (Merck, 115 930, Germany) in 1% sodium borate (Panreac, 141 644, Spain). The last semithin section (which corresponds to the section immediately adjacent to the block surface) was examined under light microscope and photographed to accurately locate the neuropil regions to be examined (Fig. 2 ).
7
Transmission Electron Microscopy of ER
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BMDCs were placed into 2.5% glutaraldehyde (Sigma-Aldrich, G6257) for at least 2 h in 4°C. The cells were post-fixed in 1% osmium tetroxide (Sigma-Aldrich, 201,030), dehydrated in acetone (Sigma-Aldrich, 650,501) and embedded in epoxy resin (Sigma-Aldrich, O5500). Resin-embedded blocks were cut into 50–60 nm ultrathin sections. The ultrathin sections were stained with both uranyl acetate and lead citrate. The changes in the endoplasmic reticulum were examined with a transmission electron microscope.
8
TEM Fixation and Embedding of Oocytes
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For TEM inspection, oocytes were fixed in 3% glutaraldehyde (G5882, Sigma Aldrich) in 0.1M cacodylate buffer (C0250, Sigma Aldrich) (pH 7.2-7.8) at room temperature overnight. Then, oocytes were postfixed with 1% osmium tetroxide (O5500, Sigma Aldrich) in 0.1 M sodium cacodylate buffer supplemented with 1.5% potassium ferrocyanide (1049821000, Sigma Aldrich) for 1 hour. Individual samples were mounted into agarose, dehydrated, and then embedded in epoxy resin, as previously described (24 24. Trebichalská, Z.
9
Transmission Electron Microscopy Tissue Preparation
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EC sections were post-fixed for 24h in a solution containing 2% paraformaldehyde, 2.5% glutaraldehyde (TAAB, G002, UK) and 0.003% CaCl 2 (Sigma, C-2661-500G, Germany) in sodium cacodylate (Sigma, C0250-500G, Germany) buffer (0.1M). These sections were washed in sodium cacodylate buffer (0.1M) and treated with 1% OsO 4 (Sigma, O5500, Germany), 0.1% potassium ferrocyanide (Probus, 23345, Spain) and 0.003% CaCl 2 in sodium cacodylate buffer (0.1M) for 1h at room temperature. After (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission.
washing in PB, sections were stained with 2% uranyl acetate (EMS, 8473, USA), and then dehydrated and flat-embedded in Araldite (TAAB, E021, UK) for 48h at 60ºC (DeFelipe and Fairén, 1993). Embedded sections were glued onto a blank Araldite block and trimmed. Semithin sections (1-2 μm thick) were obtained from the surface of the block and stained with 1% toluidine blue (Merck, 115930, Germany) in 1% sodium borate (Panreac, 141644, Spain). The last semithin section (which corresponds to the section immediately adjacent to the block surface) was examined under light microscope and photographed to accurately locate the neuropil regions to be examined (Fig. 2).
washing in PB, sections were stained with 2% uranyl acetate (EMS, 8473, USA), and then dehydrated and flat-embedded in Araldite (TAAB, E021, UK) for 48h at 60ºC (DeFelipe and Fairén, 1993). Embedded sections were glued onto a blank Araldite block and trimmed. Semithin sections (1-2 μm thick) were obtained from the surface of the block and stained with 1% toluidine blue (Merck, 115930, Germany) in 1% sodium borate (Panreac, 141644, Spain). The last semithin section (which corresponds to the section immediately adjacent to the block surface) was examined under light microscope and photographed to accurately locate the neuropil regions to be examined (Fig. 2).
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