Mouse eyes fixed in 3% glutaraldehyde and 1% paraformaldehyde in 0.08 M sodium cacodylate buffer, pH 7.4, were sequentially en bloc stained with 1% osmium 1.5% potassium ferricyanide in 0.08M cacodylate buffer, 1% aqueous sodium thiocarbohydrazide, 1% aqueous osmium, 1% aqueous uranyl acetate followed by Walton’s lead aspartate (50 (link)). The samples were dehydrated using a series of increasing concentrations of ethanol (50, 70, 90, 3x 100%) followed by propylene oxide and infiltration in a mixture of propylene oxide and Durcupan ACM resin (1:1), before embedding in Durcupan ACM resin. Blocks cut from the embedded specimens were superglued onto aluminum pins prior to coating with gold palladium. Images were acquired in between sequential sectioning (100nm thick) of the block surface using a Gatan 3View system (Gatan Inc, Abingdon, UK) and a Zeiss Sigma VP field emission scanning electron microscope (Zeiss, Cambridge, UK). The images were re-aligned using the StackReg plugin (EPFL) in ImageJ (NIH) and modelling was performed using Amira 5.3.3 software (FEI). Measurements of photoreceptor inner segment ER and mitochondria of were made using 9 and 12 rods respectively from Reep6 KO and heterozygous mice in Amira and analyzed in Prism (Graphpad).
Sigma vp field emission scanning electron microscope
Manufactured by Zeiss
Sourced in Germany, United Kingdom
The Sigma VP Field Emission Scanning Electron Microscope is a high-performance imaging and analysis tool designed for a wide range of applications. It features a field emission electron source, which provides high-resolution imaging capabilities. The microscope operates under variable pressure conditions, allowing for the analysis of non-conductive and outgassing samples without the need for extensive sample preparation.
Automatically generated - may contain errors
Lab products found in correlation
Hexamethyldisilazane, by Merck Group (1 mentions)
Sputter coater 108 auto, by Cressington (1 mentions)
Dsa100, by Krüss (1 mentions)
Bx3 25nd25, by Olympus (1 mentions)
Zetasizer nano z, by Malvern Panalytical (1 mentions)
Avance 3 400, by Bruker (1 mentions)
Szx10 microscope, by Olympus (1 mentions)
Icon3, by Bruker (1 mentions)
22 protocols using sigma vp field emission scanning electron microscope
1
Ultrastructural Analysis of Photoreceptor Organelles
2
SEM Imaging of Contact Lenses
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In a subsequent experiment, following overnight incubation at 37 °C, the soft contact lenses were fixed in 2.5% glutaraldehyde/0.1 M cacodylate buffer pH 7.4. The lenses were processed in the Electron Microscopy Core at the University of Texas Southwestern Medical Center according to the following protocol. Briefly, the lenses were washed in 0.1 M cacodylate pH 7.4 and then subjected to secondary fixation using 1% osmium. After rinsing with water, the lenses underwent an ethanol dehydration series followed by drying with hexamethyldisilazane (Sigma-Aldrich). Lenses were air dried, mounted on aluminum stubs, and sputter-coated with gold-palladium in a Cressington 108 Auto Sputter Coater (Cressington Scientific Instruments, Watford, UK). Lenses were then imaged with a Zeiss Sigma VP field emission scanning electron microscope (Zeiss, Jena, Germany).
3
Fabrication of Epoxy-based Monolithic Filters
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The polymerization of epoxy-based monoliths was already published elsewhere [20 (link)], and the monolithic filter discs used in this work were produced with adjustments after published protocols [38 (link),39 (link)]. In short, polytetrafluoroethylene molds with 16.3 × 60.0 mm of internal diameter were used (Supplementary Figure S1a ) to obtain monolithic filter columns that were cut into discs with height of either 3 or 10 mm (Supplementary Figure S1c,d ). The porogenic reaction mixture consisting of toluene and tert-butyl methyl ether (60:40, v/v) was heated to 29 °C. Subsequently, the initiator boron trifluoride diethyl etherate (BF3·Et2O) in 1,4-dioxane (1:10, v/v) was added to a concentration of 1.25%, and the components were mixed thoroughly. Then, the monomer polyglycerol-3-glycidyl ether (monomer/porogenic mixture ratio 20:80, v/v) was added, and after vigorous mixing, the solution was filled into the molds and incubated for 45 min at 29 °C (Supplementary Figure S1b ). Afterward, the resulting monolithic columns were removed from their molds, stored in methanol overnight and air-dried. Scanning electron microscopy of monolithic filters was performed on a ZEISS SIGMA VP Field Emission Scanning Electron Microscope (Carl Zeiss Microscopy GmbH, Jena, Germany) as described elsewhere in detail [31 (link)].
4
Corneal Ultrastructural Analysis Protocol
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Following either debridement of the limbal and corneal epithelium, or debridement as well as treatment with 1M N-heptanol for 120s, corneas were fixed overnight at 4°C in Karnovsky's medium, containing 2.5% gluteraldehyde, 1% paraformaldehyde, 80mM cacodylate buffer, and 20mM NaOH. Samples were washed three times for 5 minutes each with a wash buffer containing 1M sodium cacodylate. A 2% aqueous osmium tetroxide treatment was performed for 2 hours, followed by five 5-minute washes with ddH20. Samples were then dried serially at room temperature with 50%, 70%, 90% ethanol for 15 minutes each, and then 100% ethanol and 100% methanol twice each for 30 minutes, and finally 100% Hexamethyldisilazane once for 30 minutes, and once more overnight. At room temperature, 100% conductive silver paint was applied and left to dry overnight. Finally, gold/palladium sputter coating was applied to 1.5-1.7nM (Cressington 108 auto sputter coater, Watford, UK). Samples were imaged using a Zeiss Sigma VP field emission scanning electron microscope (Carl Zeiss, Hertfordshire, UK).
5
Measuring Cell-Nanotopography Interactions
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Nanotopograph
arrangements were subsequently measured by SEM (FEI Nova, under 10
kV, WD = 10 mm) after sputter-coated with 3 nm Au. For cell SEM imaging,
HOb cells were seeded on the nanotopographies at 2000 cells/cm2 and were fixed in 1.5% glutaraldehyde/0.1 M sodium cacodylate
buffer for 1 h at 4 °C. After fixation, cells were then washed
three times in 0.1 M sodium cacodylate buffer before incubation in
1% osmium tetroxide/0.1 M sodium cacodylate buffer. Afterward, nanotopographies
were washed three times with deionized (DI) water and stained with
0.5% uranyl acetate/distilled water for 1 h in the dark, and followed
by the dehydration procedure through an ethanol gradient (30, 50,
70, 90, and 100% ethanol). Samples were loaded onto a critical point
dryer (liquid CO2) for 1 h 30 min and then given a gold/palladium
coating using a POLARON SC515 SEM COATER. High-resolution secondary
electron of cell nanotopography interactions imaging was performed
at the Imaging, Spectroscopy and Analysis Centre (ISAAC) at the University
of Glasgow, UK. The images were acquired with a Zeiss Sigma VP Field
Emission scanning electron microscope under high vacuum conditions
using 5 kV accelerating voltage, an aperture size of 30 mm, and at
a working distance of 5 mm.
arrangements were subsequently measured by SEM (FEI Nova, under 10
kV, WD = 10 mm) after sputter-coated with 3 nm Au. For cell SEM imaging,
HOb cells were seeded on the nanotopographies at 2000 cells/cm2 and were fixed in 1.5% glutaraldehyde/0.1 M sodium cacodylate
buffer for 1 h at 4 °C. After fixation, cells were then washed
three times in 0.1 M sodium cacodylate buffer before incubation in
1% osmium tetroxide/0.1 M sodium cacodylate buffer. Afterward, nanotopographies
were washed three times with deionized (DI) water and stained with
0.5% uranyl acetate/distilled water for 1 h in the dark, and followed
by the dehydration procedure through an ethanol gradient (30, 50,
70, 90, and 100% ethanol). Samples were loaded onto a critical point
dryer (liquid CO2) for 1 h 30 min and then given a gold/palladium
coating using a POLARON SC515 SEM COATER. High-resolution secondary
electron of cell nanotopography interactions imaging was performed
at the Imaging, Spectroscopy and Analysis Centre (ISAAC) at the University
of Glasgow, UK. The images were acquired with a Zeiss Sigma VP Field
Emission scanning electron microscope under high vacuum conditions
using 5 kV accelerating voltage, an aperture size of 30 mm, and at
a working distance of 5 mm.
6
Nanoparticle Characterization via SEM
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A Sigma VP field emission scanning electron microscope (Carl-Zeiss AG) was used to obtain the particle images. The microscope was operated with the InLens detector at a 6 kV acceleration voltage. 5 µL of NP dispersions were pipetted on mica substrates and air-dried. Before the measurement, samples were coated with a thin layer of platinum (4 nm) via sputter coating (CCU-010 HV, Safematic). ImageJ was used to estimate the NP size from 300 and 500 particles per image, for Ace-DEX and PLGA NPs, respectively.
7
Scanning Electron Microscopy of Nanomaterials
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Imaging of the particles was performed using a Sigma VP Field Emission Scanning Electron Microscope (Carl-Zeiss, Jena, Germany) equipped with an Inlens detector and operated at an accelerating voltage of 5 to 6 kV. Prior to imaging, the samples were drop-casted on a mica plate and coated with a 4 nm platinum layer using a CCU-010 HV sputter (Safematic, Zizers, Switzerland).
8
Characterizing NP Surface Morphology via SEM
9
Nanoparticle Surface Morphology Analysis
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Surface morphology of the nanoparticles was studied using scanning electron microscopy. Lyophilized nanoparticles were imaged using a Sigma VP Field Emission Scanning Electron Microscope (Carl Zeiss Microscopy Ltd, NY, USA). Cressington 108 Auto Sputter Coater for 30 s was utilized prior to imaging. The target used for coating was gold/platinum.
10
Soil Microstructural Analysis via SEM
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After UCS tests, intact chunks of stabilized soil were subjected to SEM imaging to observe the microstructure of the samples. The samples were dried in the oven at 35 °C for 24 h. Then, they were attached onto a SEM mount by carbon conductive tabs. After ensuring sufficient electric grounding using carbon paint, samples were coated with a conductive tape. A Zeiss Sigma VP Field Emission Scanning Electron Microscope (Oberkochen, Germany) was used for specimen observation.
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