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Identifying Nanotoxicity at the Cellular Level Using Electron Microscopy
Published in Suresh C. Pillai, Yvonne Lang, Toxicity of Nanomaterials, 2019
Kerry Thompson, Alanna Stanley, Emma McDermott, Alexander Black, Peter Dockery
To provide sufficient contrast to the sample, heavy metal salt staining is required. Unlike in light microscopy where different cellular components can be selectively stained, for electron microscopy stains are compounds or ions of high atomic number which scatter the electron beam and are only semi-selective. However, the interaction of such stains with certain chemical groups is known and should be considered depending on the nanoparticle material as they may not only enhance the contrast of the nanoparticles, but by attaching to them may increase their size. The standard TEM counterstains for biological samples are uranyl acetate followed by lead citrate. Uranyl acetate interacts with anionic compounds and binds strongly to phosphate groups, particularly those of nucleic acids and membrane phospholipids, in addition to sialic acid carboxyl groups of proteins and lipids. Lead citrate acts as a mordant for osmium tetroxide and uranyl acetate and therefore further enhances the contrast provided by them. It also binds to negatively charged amino acids of proteins, hydroxyl groups of carbohydrates, and phosphate groups of nucleic acids (Bozzola, 2014b).
Anticancer activity of vanadium nanoparticles against human breast cancer: an in vitro study
Published in Inorganic and Nano-Metal Chemistry, 2023
Canan Vejselova Sezer, Hatice Mehtap Kutlu
MCF-7 (ATCC® HTB-22™) human breast cancer cells were obtained from the American Type Culture Collection (Manassas, USA). Vanadium (IV)-oxid sulfate pentahydrate pure (VOSO4) purchased from (Riedel-de Haen cat: 14229 Lot: s29267-275, CA) was used as a test agent in the experimentations. Dimethyl sulfoxide (DMSO), MTT (3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyl-2H-tetrazolium bromide) (M2003), Dulbecco’s Phosphate Buffered Saline (PBS) were purchased from Sigma-Aldrich (St. Louis, USA), Roswell Park Memorial Institute medium (RPMI-1640) was obtained from GIBCO (Grand Island, USA), fetal bovine serum (FBS) and penicillin-streptomycin were from Merck Schuchardt (Darmstadt, Germany). Osmium tetraoxide, glutaraldehyde, epoxy, propilen oxide, uranyl acetate, lead citrate were obtained from Electron Microscopy Science (USA). Annexin-V apoptosis detection kit was from BD, Pharmingen (USA) and phalloidine, Anti-E cadherin were obtained from Thermo Scientific (USA). Fluo-3, ATP, Anti-cyclin B1 and Anti-cyclin D1 were from Santa Cruz (CA, USA).
Ameliorative role of neem (Azadiracta indica) leaves ethanolic extract on testicular injury of neonatally diabetic rats induced by streptozotocin
Published in Egyptian Journal of Basic and Applied Sciences, 2020
Abd El-Fattah B. M. El-Beltagy, Amoura M. Abou El-Naga, El-Sayed M. El-Habibi, Sara El-Said M. Shams
For all examined groups, small pieces of testes were fixed in 5% buffered glutaraldehyde (pH 7.3) at 4°C. After 48 h, postfixation the pieces of testes were washed in 5% buffered sucrose (pH 7.3). The pieces of testes were cut into fine blokes less than 1 mm2 and then fixed in 1% osmium tetroxide for 2 hr followed by washing twice in phosphate buffer (pH 7.3) for 10–15 min [23]. After that, the sections were dehydrated in ascending series of alcohol, propylene oxide and then embedded in Araldite. The semi-thin sections (0.6–0.7 µm thick) were cut with aid of glass knives on ultra-microtome. The obtained sections were stained with toluidine blue and examined by a binocular light microscope. According to the investigated semi-thin sections, the ultrathin sections were cut at 600–700A° and collected on cleaned copper grids. The prepared grids were stained in uranyl acetate and lead citrate. Finally, the grids were investigated and photographed in a JEOL 1200 EXIL TEM in EM center of Mansoura University, Egypt.
The role of thioredoxin and glutathione systems in arsenic-induced liver injury in rats under glutathione depletion
Published in International Journal of Environmental Health Research, 2022
Yuanyuan Li, Kun Liang, Lin Yuan, Jing Gao, Linquan Wei, Lijun Zhao
Approximately 0.5 to 1 mm3 of fresh liver tissue was quickly inserted in a tiny tube containing 2 to 3 mL of precooled fixing solution (2.5% glutaraldehyde) for 2 h at 4°C. The samples were then serially dehydrated with 50%, 70%, 90%, and 100% ethanol. The samples were soaked in a mixture of pure acetone and embedding agent (2:1, v/v) at room temperature for 2 h, followed by a 3 h soaking in a mixture of pure acetone and embedding agent (1:2, v/v) at room temperature. The samples were encased in epoxy resin overnight at 37°C and then for 48 hours at 60°C. Then, the samples were sliced and stained with lead citrate and uranyl acetate. The materials were then examined using H-7650 80 kV transmission electron microscope (HITACHI, Japan).