The morphology of the produced Ag-HA nanocomposite was compared to that of virgin HA using the scanning electron microscope (SEM) JEOL JEM 1010 equipment (JEOL, Tokyo, Japan), equipped with an EDAX detector (X-act, Oxford instruments) was used for elemental mapping. The study conducted TEM measurements using a JEOL JEM 1010 instrument with an acceleration voltage of 100 kV. The preparation of samples for TEM analysis involved depositing a diluted suspension of the sample onto an ultrathin carbon-coated copper grid. The study utilized a Siemens D5000 powder X-ray diffractometer to collect X-ray diffraction (XRD) patterns within the range of 2 h = 5–100, using CuKa radiation (λ = 1.54056 Å). These patterns were then compared to the crystallographic information files (CIF) obtained from the crystallographic open data base (COD). The analysis of chemical bonding in Ag-HA nanocomposite was carried out using an X-ray photoelectron spectroscopy XPS/ESCA equipment. Raman spectra of Ag-HA nancomposite was conducted via Renishaw in Via Reflex Raman spectrometer (Renishaw, Gloucestershire, UK). The FTIR spectra were examined using a JASCO FTIR 3600 spectrometer. Agilent Cary 60 UV–Vis spectrophotometer was used to study UV–Vis spectra.
Jem 1010
Manufactured by JEOL
Sourced in Japan, United States, Germany, Spain, China
The JEM-1010 is a transmission electron microscope (TEM) manufactured by JEOL. It is designed for high-resolution imaging of a wide range of samples. The JEM-1010 provides a maximum accelerating voltage of 100 kV and offers a high level of stability and reliability for routine imaging applications.
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Lab products found in correlation
Ultracut uct, by Leica (17 mentions)
Glutaraldehyde, by Merck Group (11 mentions)
Em uc7, by Leica (11 mentions)
Sc1000, by Ametek (10 mentions)
Embed 812 resin, by Electron Microscopy Sciences (8 mentions)
Paraformaldehyde (pfa), by Merck Group (8 mentions)
Zetasizer nano z, by Malvern Panalytical (7 mentions)
Diamond knife, by Electron Microscopy Sciences (7 mentions)
S 4800, by Hitachi (7 mentions)
Ultramicrotome, by Leica (7 mentions)
Poly bed 812, by Polysciences (6 mentions)
Ultracut uct ultramicrotome, by Leica (6 mentions)
Fe sem s 4800, by Hitachi (5 mentions)
Ab134045, by Abcam (5 mentions)
Temcam f416, by TVIPS (5 mentions)
Api zym, by bioMérieux (5 mentions)
Uc7 ultramicrotome, by Leica (4 mentions)
Uc6 ultramicrotome, by Leica (4 mentions)
Ultramicrotome, by Reichert Technologies (4 mentions)
Epon 812 resin, by TAAB (4 mentions)
752 protocols using jem 1010
1
Comprehensive Characterization of Ag-HA Nanocomposite
2
Chloroplast Ultrastructure Observation
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3
Negatively Stained Vesicle Observation and QFDE-EM Analysis
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For observation of negatively stained vesicles, samples were placed on Cu400 mesh grids (JEOL, Tokyo, Japan) that were pretreated with 0.01% α-poly-L -lysine. Bacterial cells and vesicles were stained with 2% (NH4)6Mo7O24 and observed using a JEM-1010 (JEOL) at 80 kV that was equipped with a FastScan-F214 (T) CCD camera (TVIPS, Gauting, Germany).
For quick freeze and replica electron microscopic (QFDE-EM) observations, the protocol was as described previously (Takaki et al., 2020 (link)). Briefly, bacterial cells were centrifuged, washed, mixed with a rabbit lung slab and mica flakes, and then placed on a paper disk attached to an aluminum disc. The samples were quickly frozen in liquid helium using a CryoPress (Valiant Instruments, St. Louis, MO, United States). The specimens were placed in a chamber maintained at −180°C using a JFDV freeze-etching device (JEOL). The samples were freeze-fractured with a knife and freeze-etched. Subsequently, the samples were coated with platinum and then coated with carbon. The replicas were floated in full-strength hydrofluoric acid, rinsed in water, cleaned with a commercial bleach containing sodium hypochlorite, and rinsed in water. Replica specimens were placed onto grids and observed using a JEM-1010 (JEOL).
For quick freeze and replica electron microscopic (QFDE-EM) observations, the protocol was as described previously (Takaki et al., 2020 (link)). Briefly, bacterial cells were centrifuged, washed, mixed with a rabbit lung slab and mica flakes, and then placed on a paper disk attached to an aluminum disc. The samples were quickly frozen in liquid helium using a CryoPress (Valiant Instruments, St. Louis, MO, United States). The specimens were placed in a chamber maintained at −180°C using a JFDV freeze-etching device (JEOL). The samples were freeze-fractured with a knife and freeze-etched. Subsequently, the samples were coated with platinum and then coated with carbon. The replicas were floated in full-strength hydrofluoric acid, rinsed in water, cleaned with a commercial bleach containing sodium hypochlorite, and rinsed in water. Replica specimens were placed onto grids and observed using a JEM-1010 (JEOL).
4
Vesicle and Cell Volume Analysis
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TEM sections of hCMEC/D3 and ciGENC cells were used to analyse vesicular diameter and cell volume. Each section corresponded to a well with a single treatment. As each treatment had 3 wells, one was randomly picked for this analysis and 3 independent experiments were analysed per cell type, resulting in 3 sections analysed altogether. At 20,000x magnification on JEM 1010 (Jeol, Japan), about 100 vesicular diameters were measured per section in cells that were randomly picked. The data set was analysed with Microsoft Office Excel 2010 and GraphPad Prism. Similarly, to obtain volumes of brain cells in comparison to kidney cells, 3 independent experiments were analysed for each cell type. At magnification of 2,000X on JEM 1010 (Jeol, Japan) 10 images per treatment were taken randomly. The cell area was measured using ImageJ. The data were analysed as a distribution histogram and means of each independent experiment were calculated and analysed by GraphPad Prism.
5
Transmission Electron Microscopy of Brucella OMVs
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Twenty-five microliters from the OMVs suspension (25 μg of protein) from both strains were placed onto copper grids coated with formvar and they were subsequently dried using filter paper. Then, 1% phosphotungstic acid was added onto the samples. The grids were allowed to dry for 10 h and they were observed with the aid of a transmission electron microscope (JEOL model JEM 10-10). In order to observe OMVs released from whole bacteria, cells from the B. melitensis strain 16M and B. melitensis VTRM1 were grown on TSA plates for 36 h at 37°C. Subsequently, molten soft agar was poured onto the plates to cover the growth. Once the agar had solidified, small agar cubes (2 mm) were cut. Blocks were fixed using 2.5% glutaraldehyde in PBS, rinsed with Sorensen’s PBS, dehydrated with ethanol, and prepared for transmission electron microscopy (TEM). Thin-section preparations were stained with OsO4, and they were observed using the JEOL model JEM 10–10, transmission electron microscope. Images were obtained using the aforementioned transmission electron microscope at ENCB’s (IPN, Mexico City, Mexico) Microcopy Facility.
6
Ultrastructural Analysis of Brucella Membrane Blebs
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Thirty microliter of membrane blebs (25 μg of protein) from both strains were placed onto copper grids coated with formvar and dried using filter paper. Then, 1% phosphotungstic acid was added on the samples. The grids were allowed to dry 10 h and they were observed in a transmission electron microscope (JEOL model JEM 10-10). The micrographs were taken with ATM image capture engine V. 5. 4. 2 software with different magnifications. The membrane blebs diameter was measured and the blebs were counted from ten fields with the same software. To observe membrane blebs released from the whole bacteria, cells of B. abortus strain 2308 and B. abortus strain RB51were grown on TSA plates for 36 h at 37°C, and then molten soft agar was poured to cover the growth. Once solidified the agar, small cubes (2 mm) of agar were cut. Blocks were fixed in 2.5% glutaraldehyde in PBS, rinsed with Sorensen’s PBS, dehydrated with ethanol, and prepared for transmission electron microscopy (TEM). All preparations were stained with OsO4, and they were observed in a transmission electron microscope (JEOL model JEM 10-10) and the micrographs were obtained as described above.
7
Visualizing Bacterial Outer Membrane Vesicles
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Twenty microliter of purified OMVs (25 μg of protein) were placed onto copper grids coated with formvar and dried using filter paper. 1% phosphotungstic acid was added and the grids were allowed to stand overnight at room temperature; they were observed under the transmission electron microscope (JEOL model JEM 10-10).
To observe vesicles released from bacteria, A. hydrophila ATCC® 7966TM was grown in TSA plates overnight, and then molten soft agar was poured over growth. Once solidified the agar, small cubes (2 mm) of agar were cut. Blocks were fixed in glutaraldehyde 2.5% in PBS, rinsed with Sorensen’s PBS, dehydrated with ethanol, and prepared to transmission electron microscopy (TEM). All preparations were stained with OsO4, and they were observed in transmission electron microscope (JEOL model JEM 10-10).
To observe vesicles released from bacteria, A. hydrophila ATCC® 7966TM was grown in TSA plates overnight, and then molten soft agar was poured over growth. Once solidified the agar, small cubes (2 mm) of agar were cut. Blocks were fixed in glutaraldehyde 2.5% in PBS, rinsed with Sorensen’s PBS, dehydrated with ethanol, and prepared to transmission electron microscopy (TEM). All preparations were stained with OsO4, and they were observed in transmission electron microscope (JEOL model JEM 10-10).
8
Ultrastructural Analysis of SLO-Exposed Trophozoites
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Trophozoites that had been exposed to SLO were washed once with PBS and twice with 0.1 M sodium cacodylate buffer at 37 °C and fixed for 3 h with 2.5% glutaraldehyde in 0.2 M sodium cacodylate buffer, pH 7.4. Fixed trophozoites were washed twice with 0.1 M sodium cacodylate buffer, post-fixed with 1.0% osmium tetroxide in 0.1M sodium cacodylate at 4 °C, dehydrated with ethanol at increasing concentrations and treated with propylene oxide. Trophozoites were embedded in EmBed 812 epoxy resins, polymerized blocks were cut using an ultramicrotome, and thin sections were stained with 2% uranyl acetate and 2% lead citrate. Trophozoites morphology was analyzed by transmission electron microscopy (TEM) with a JEM-1010 JEOL at 80 keV.
9
Ultrastructural Analysis of Nerve Tissue
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Nerve morphology was analyzed using Transmission electron microscopy (TEM). For this purpose, cell samples from all experimental rat groups were harvested 3 weeks after surgery and fixed overnight in 2.5% glutaraldehyde in 0.1 PBS. Samples were then fixed a second time, overnight, using 2% osmium tetroxide (TAAB Laboratories®) in PBS. Samples were then dehydrated in graded aqueous ethanol solutions and washed in propylene oxide (SERVA Electrophoresis GmbH®), finally, samples were infiltrated with epoxy resin and polymerized. The resulting blocks were cut into ultrafine sections (60 nm) using a diamond knife (Diatome®) under an LKB Ultrotome V microscope (Bromma®). Sections were then observed with a transmission electron microscope (JEM-1010 JEOL®) using a double contrast method (uranyl and plumb citrate). Images of 10 random fields were captured foreach section and hystomorphometric analysis was performed using NIS Elements microscope imaging software (Nikon®).
10
Comprehensive Particle Characterization Protocol
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UV-Vis-NIR spectra of the particles were obtained using quartz cuvette with a 1-cm light path using a UV-Vis spectrophotometer (Beckman Coulter, Fullerton, CA, USA). The images of particles during synthesis and modification were captured under transmission electron microscope (JEM 1010 JEOL, Tokyo, Japan) by placing aliquots of aqueous particle suspension on carbon-enhanced copper grids and accelerating the machine at 200 kV. The size distribution of the particles was analyzed using a Zeta instrument (ELS-8000, OTSUKA Electronics Co. Ltd., Japan). The X-ray diffraction (XRD) spectra of the particles were measured using XRD (X’Pert-MPD, Philips, Amsterdam, The Netherlands) with Cu-Kα radiation. To analyze the functional groups of the particles at different stages, aliquots of freeze-dried product were mixed with KBr and pelletized for FTIR analysis (Perkin Elmer Inc., Waltham, Massachusetts, USA) at a resolution of 4 cm−1 over the wavelength range 500 to 4000 cm−1. Thermogravimetric analysis was performed using TGA 7, Pyris 1 (Perkin Elmer, MA, USA).
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