For the observation of protein particles, the purified BoCesA was diluted to 0.06 mg/mL. For the synthesized cellulose, the 1 mL samples were first centrifuged at 10,000×g at 4 °C for 30 min and the upper 900 µL solution was discarded. 5 µL sample solution from the remaining liquid was dropped onto a discharged carbon film-coated 300 mesh copper grid (CF300-Cu, Electron Microscopy Sciences) and stained with 2% uranyl acetate. The TEM images were taken by a Hitachi H-7650 TEM (75 kV, 30–120 K magnification, Erlangshen ES500W Model 782 CCD camera).
Cf300 cu
Manufactured by Electron Microscopy Sciences
Sourced in United States
The CF300-CU is a compact and versatile field emission scanning electron microscope (FESEM) designed for high-resolution imaging of a wide range of samples. It features a cold field emission gun, which provides a bright, stable, and low-energy electron beam for high-quality imaging. The CF300-CU is equipped with essential imaging modes, including secondary electron and backscattered electron detection, enabling users to observe surface topography and material contrast with high resolution.
Automatically generated - may contain errors
Lab products found in correlation
Whatman, by Cytiva (6 mentions)
H 7650 tem, by Hitachi (5 mentions)
Morgagni 268 microscope, by Thermo Fisher Scientific (4 mentions)
Whatman filter paper, by Cytiva (3 mentions)
Zetasizer nano z, by Malvern Panalytical (2 mentions)
Vwr signature forced air safety ovens, by Avantor (2 mentions)
Glutaraldehyde, by Electron Microscopy Sciences (2 mentions)
Uranyl formate, by Electron Microscopy Sciences (2 mentions)
Em420 electron microscope, by Philips (2 mentions)
Zetasizer nano zs zen3600, by Malvern Panalytical (2 mentions)
Talos l120c transmission electron microscope, by Thermo Fisher Scientific (2 mentions)
Jem 1400 transmission electron microscope, by JEOL (1 mentions)
Spectramax id5 plate reader, by Molecular Devices (1 mentions)
Tecnai spirit tem, by Thermo Fisher Scientific (1 mentions)
Jem 100cx 2 electron microscope, by JEOL (1 mentions)
Em grid, by Electron Microscopy Sciences (1 mentions)
Jem 1010 transmission electron microscope, by JEOL (1 mentions)
Tla120 rotor, by Beckman Coulter (1 mentions)
Tecnai f20 tem, by Thermo Fisher Scientific (1 mentions)
Ace600 coating device, by Leica (1 mentions)
38 protocols using cf300 cu
1
Protein Particle Observation via TEM
2
Characterization of Nanoparticles by TEM
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Transmission electron microscopy (TEM) was performed on a JEOL JEM 1400 Transmission Electron Microscope at 100 kV. The sample was prepared as follows: 5 µL of ~0.1 nM nanoparticles solution was left to adsorb onto a PEI-covered TEM grid (CF300-Cu, Electron Microscopy Sciences) for 30 min. The remaining solution was washed off with water and the dry grids were used for imaging. Zeta potential and hydrodynamic size measurements were performed using Zetasizer Nano ZS (Malvern) on particles dispersed in 10 mM HEPES buffer (pH 7.2), (Table S1 , Supporting Information ). Nanoparticle tracking analysis (NTA) NS500 (Malvern) was used to determine nanoparticle concentration. The concentration of gold seeds was calculated from the absorption spectrum. SpectraMax ID 5 plate reader (Molecular Devices) was used for optical characterization of nanoparticles (Figure S2 ).
3
Cdc19 Structural Analysis by TEM
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After thawing on ice, Cdc19 samples were cleared by centrifugation (21000 g, 4 °C, 10 min), diluted to 0.2 mg/ml in purification buffer pH 7.4, and the pH was adjusted to 6-7. Subsequently, a non-glow discharged carbon film 300 mesh copper grid (CF300-CU, Electron Microscopy Sciences) was submerged in this Cdc19 dilution and incubated for 10 min at 42 °C. Excess sample was then manually removed with Whatman filter paper. Where indicated, grids with pre-formed Cdc19 aggregates were incubated for 20 min with FBP or F6P dissolved in purification buffer to the indicated concentration, or simply washed twice with purification buffer, and stained with two drops of 2% uranyl acetate. TEM micrographs were acquired with a FEI Morgagni 268 microscope at 100 kV using a CCD 1376 × 1032 pixel camera at different magnifications.
4
Cryo-TEM Imaging of ELP-CLP Conjugates
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Samples for TEM were prepared on carbon-coated copper grids (CF300-Cu, Electron Microscopy Sciences Inc.). ELP-CLP conjugates were dissolved in water at concentration of 1 mg/ml [consistent with DLS experiments and to prevent precipitation of the phosphotungstic acid (PTA) stain]. The samples were agitated before sample preparation. The sample solution (5 μl) was drop casted on the grid and blotted after 60 s. For staining, 1% PTA (pH adjusted to 7.0 using 1 M NaOH) as a negative stain was used. The grids, pipette tips, and samples were incubated in an isothermal oven (VWR Signature Forced Air Safety Ovens, VWR Inc.) at desired temperature (4°, 25°, 37°, 50°, and 80°C) for at least 1 hour before sample preparation, which was also conducted in the oven. The PTA solution (3 μl) was drop casted on the grid and blotted after 10 s. The samples were dried in the oven at the desired temperature for 1 hour and then were air-dried for more than 2 hours. TEM images were taken on a 2.1 TEM Tecnai 12 (JEOL USA Inc., Peabody, MA) at an acceleration voltage of 120 keV. Each ELP-CLP conjugate was produced and characterized by three separate times, indicating the repeatable for each ELP-CLP conjugate.
5
Transmission Electron Microscopy Analysis of hBM-MSC-sEVs
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For TEM analysis, hBM-MSC-sEVs were isolated as described in the “hBM-MSC-sEV isolation ” section from a starting volume of 15 mL of CCM. Following hBM-MSC-sEV isolation, the pellet was suspended in 300 μL of filtered PBS which was then filtered using vivaspin 300 kDa filters (Satorius, Cat#VS0651), following a rinse of the filters with 200 μL of filtered PBS centrifuged at 2000×g for 3 min. The concentrated hBM-MSC-sEVs were suspended in an equal volume of 4% PFA for 30 min. Next, two 50 μL drops of hBM-MSC-sEV/PFA suspension were deposited on parafilm and on which carbon-coated electron microscopy grids (Electron Microscopy Sciences, Cat#CF300-CU) were inverted and placed for 5 min on each 50 μL sample followed by a rinse with 50 μL drops of PBS on a sheet of parafilm. Grids were blotted on filter paper to remove excess and let dry for an hour before imaging. The TEM imaging was performed using a FEI Tecnai Spirit TEM with a LaB6 emitter, operating at 120 kV. The images were acquired with an Eagle camera with 4 k × 4 k resolution.
6
Assembling M1, NM1, and M1-V97K Oligomers
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Purified full-length M1 was diluted into cold 50 mM HEPES (pH 7) to a final concentration of 1.8 μM, assembled by the addition of cold 2 M NaCl at 4°C for 2–24 h, and applied onto glow-discharged, carbon-coated, 300 mesh copper EM grids (CF300-Cu; Electron Microscopy Sciences, Hatfield, PA, USA). Purified NM1 was diluted into cold 50 mM HEPES (pH 7) to a concentration of 170 μM and assembled by the addition of cold 2 M NaCl at 4°C overnight. M1-V97K oligomers were assembled by diluting protein stock into 50 mM HEPES (pH 7) to a final concentration of 54 μM and assembled by the addition of 3 M NaCl at 37°C overnight.
7
Negative-Stain Electron Microscopy Sample Preparation
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Samples were diluted to 100 μg/ml in 20 mM HEPES pH 7.4, 150 mM NaCl, 5% glycerol, 7.5 mM glutaraldehyde (Electron Microscopy Sciences) and incubated for 5 min before quenching the glutaraldehyde by the addition of 1 M Tris (to a final concentration of 75 mM) and 5 min incubation. A 5-μl drop of sample was applied to a glow-discharged carbon-coated grid (Electron Microscopy Sciences, CF300-Cu) for 10 to 15 s, blotted, stained with 2% uranyl formate (Electron Microscopy Sciences), blotted, and air-dried. Images were obtained using a Philips EM420 electron microscope at 120 kV, 82,000× magnification, and a 4.02 Å pixel size. RELION (57 (link)) software was used for particle picking and 2D and 3D class averaging.
8
Liposomal Nanoparticle Characterization by TEM
9
Visualization and Analysis of Drp1-Actin Interactions
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Drp1 was diluted in 50 mM KCl, 1 mM MgCl2, 1 mM EGTA, 10 mM Hepes pH 7.4. To remove potential aggregates or small Drp1 nuclei, Drp1 was centrifuged at 100,000 rpm for 20 min at 4°C in a TLA-120 rotor (Beckman), and the supernatant was stored on ice for use. 4 μM actin was polymerized for 7 min in G-buffer plus 50 mM KCl, 1 mM MgCl2, 1 mM EGTA, 10 mM Hepes pH 7.4. Drp1 was added to polymerized actin for a final concentration of 1.3 μM Drp1 and 2 μM actin. Samples (30 μl) were absorbed onto EM grids (Electron Microscopy Sciences, CF300-Cu) for 4 min, then blotted gently with filter paper. Grids were subsequently stained with 1% uranyl acetate solution for 1 min, and again blotted gently with filter paper. The prepared grids were imaged on a JEOL JEM 1010 transmission electron microscope operated at 100 keV acceleration. Images were collected using an XR-41B AMT digital camera and capture engine software (AMTV 540; Advanced Microscopy Techniques). Filament widths were quantified using ImageJ.
10
Transmission Electron Microscopy of Tellurium Nanostructures
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Te-nanostructure extracts were recovered from BCP1 resting cells following the procedure published in our previous study21 (link), while Transmission Electron Microscopy (TEM) imaging of both TeO32−-exposed BCP1 resting cells and Te-nanostructure extracts was performed using a Hitachi H7650 TEM. For bright field (BF) and high-resolution (HR) TEM, as well as the corresponding Selected-Area Electron Diffraction (SAED) pattern of TeNRs were collected by FEI Tecnai F20 TEM at an acceleration voltage of 200 kV. The samples were prepared by mounting 5 µL of either cellular suspensions or Te-nanostructure extracts on carbon-coated copper grids (CF300-CU, Electron Microscopy Sciences), which were air-dried prior the imaging. TEM micrographs were analyzed through ImageJ software to measure the actual length of TeNRs, which was calculated considering 100 randomly chosen nanorods. The distribution was fitted to a Gaussian function to yield the average length of TeNRs.
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