Size distributions and zeta potentials of chrysin-NPs were determined by dynamic light scattering (DLS) using a Zetasizer Nano-ZS device (Malvern Instruments Ltd., Worcestershire, UK). Polydispersity index (PDI) represents a width parameter for the size-average as an intensity mean. Chrysin-NP morphology was observed by cryo-TEM (Cryo Tecnai F20; FEI Co., Hillsboro, OR, USA). Chrysin-loading efficiencies were calculated using Equation 1 . Free nonencapsulated chrysin was dissolved in dimethyl sulfoxide (DMSO) (Sigma-Aldrich) and quantified by high-performance liquid chromatography (Agilent Technologies, Inc., Santa Clara, CA, USA) and confirmed at 348 nm using a microplate reader (Infinite M200 PRO; Tecan Inc., Grödig, Austria).
Cryo tecnai f20
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Cryo Tecnai F20 is a transmission electron microscope designed for low-temperature imaging of biological samples. It features a field emission gun and a cryogenic sample stage, allowing for high-resolution imaging of specimens preserved in a frozen hydrated state.
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Lab products found in correlation
4 protocols using cryo tecnai f20
1
Characterization of Chrysin-Loaded Nanoparticles
2
Transmission Electron Microscopy Sample Preparation
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Samples were fixed with 2.5% glutaraldehyde (Sigma, USA) in 0.1 M PBS (pH 7.4) for 24 h at 4 °C. After washing with 0.1 M PBS, the samples were postfixed with 1% osmium tetroxide (OsO 4 ) in 0.1 M sodium cacodylate trihydrate buffer for 1 h at room temperature. Then, the samples were dehydrated in a series of ethanol solutions (70%-80%-85%-90%-95%-100%-100%) followed by dehydration twice in pure propylene. Samples were embedded with the Epoxy Embedding Medium Kit (Sigma, USA). The silicone membranes were detached from the PDMS chip, and the embedded samples were removed from the microchannels. The samples were then cut into ultrathin sections (70-80 nm) using an ultramicrotome (Leica, Germany) and stained with uranyl acetate and lead citrate for transmission electron microscopy (TEM) imaging. Cryo-TEM (CryoTecnai F20; FEI, USA) was used to observe the samples.
3
Quantifying Remyelinated Nerve Thickness
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To assess myelin sheath thickness, cocultures were fixed with 2% glutaraldehyde overnight at 4°C, followed by postfixation with 1% osmium tetroxide for 30 min at room temperature. After the samples had been washed three times with ultrafiltered water, they were dehydrated in an ethanol series (50, 70, 80, 85, 90, 95, and 100%). Using an ultramicrotome (Ultra Cut C; Leica, Wetzlar, Germany), samples were sectioned and mounted on copper grids. Grids were contrast stained with uranyl acetate and lead citrate. Images were captured by cryo-TEM (Tecnai F20 Cryo; FEI, Hillsboro, OR, USA). To quantitatively determine the relative thickness of individual remyelinated nerves, the g-ratio was analyzed with a g-ratio calculator plug-in for ImageJ software (available at https://imagej.nih.gov/ij/ ), which allowed semiautomated analysis of randomly selected nerve fibers (53 (link), 54 (link)). At least seven remyelinated axons per group were randomly selected for measurement, and the mean g-ratio was calculated for each group by averaging across all individual g-ratios.
4
Ultrastructural Analysis of Plant Tissues
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Freshly harvested tissues, including the primary inflorescence stems, roots, and hypocotyls, were fixed in 2.5% glutaraldehyde at 4 °C, after rinsing three times in 0.05 M cacodylate buffer (pH 6.9) containing 1% osmium tetroxide for 3 h at 4 °C. The fixed tissues were dehydrated using a graded ethanol series (30, 50, 70, 80, 90, and 100%) for 15 min at each step. The dehydrated samples were embedded in Spurr’s resin series [68 (link)] and polymerized at 60 °C for 8 h. The embedded samples were cut with a diamond knife using an Ultracut S ultramicrotome. Sections were mounted directly on 150-mesh copper grids and stained with toluidine blue. The grids were observed using a cryo-TEM (Tecnai F20 Cryo, FEI, The Netherlands), installed at the bio-imaging core facility of the Republic of Korea Institute of Science and Technology (KIST). All measurements were repeated at least three times.
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