Aliquots of 2.5–3 µl of purified PRD1 particle suspension (Table 1 ) were applied to 400 mesh R1.2/1.3 Quantifoil grids (Quantifoil Micro Tools GmbH), blotted for 2 s and immediately frozen in liquid ethane using an automated vitrification device: either a Vitrobot MarkIII (FEI) or a Cryo-Plunger 3 (Gatan). Images were taken with a 300 kV JEM3200FSC electron microscope (JEOL) equipped with in-column energy filter. A slit width of 20 eV was used for data collection. The first dataset of the virion and all procapsid data was recorded at 80 K×nominal magnification (1.42 Å/pixel sampling) with a dose of 20 e/Å2 using a Ultrascan 4000 CCD camera (Gatan) with defocus ranging from 0.5 to ∼2 µm (Table S2 ). The second dataset of virion was collected using a Ultrascan 10000 CCD camera (Gatan) binned by 2 (1.3 Å/pixel sampling) with a defocus range from 1 to 3 µm. All mutant particles were imaged on a 200 kV JEM2010F electron microscope (JEOL) with a dose of 25 e•Å−2 using a Ultrascan 4000 CCD camera (Gatan) at 40–60 k×nominal magnification sampling from 1.81 to 2.18 Å/pixel and defocus ranging from 1.5 to 3 µm (Table S2 ).
Ultrascan 4000 ccd camera
Manufactured by Ametek
Sourced in United States
The UltraScan 4000 CCD camera is a high-performance imaging device designed for scientific and industrial applications. It features a charge-coupled device (CCD) sensor that captures detailed digital images with high resolution and low noise. The camera is capable of capturing images with a resolution of up to 4096 x 4096 pixels and can operate at a variety of frame rates. The UltraScan 4000 is designed for versatile use in a range of laboratory and research settings.
Automatically generated - may contain errors
Lab products found in correlation
First light digital camera controller, by Ametek (23 mentions)
Jem 1230 transmission electron microscope, by JEOL (17 mentions)
Em uc6 ultramicrotome, by Leica (14 mentions)
Solarus 9500 plasma cleaner, by Thermo Fisher Scientific (9 mentions)
Jem 3200fsc, by JEOL (8 mentions)
3.5 1 holey carbon copper grids, by Quantifoil (8 mentions)
Tecnai f20 electron microscope, by Thermo Fisher Scientific (7 mentions)
Vitrobot mark 4, by Thermo Fisher Scientific (4 mentions)
Tecnai g2 spirit, by Ametek (3 mentions)
Digital micrograph software, by Ametek (3 mentions)
Cryoplunge 3, by Ametek (3 mentions)
Jem 2100 electron microscope, by JEOL (3 mentions)
Holey carbon copper grids, by Quantifoil (3 mentions)
Jem 3200fsc field emission microscope, by JEOL (2 mentions)
Solarus 9500 plasma cleanerand, by Thermo Fisher Scientific (2 mentions)
Mk 3 vitrobot, by Thermo Fisher Scientific (2 mentions)
Hq silver, by Nanoprobes (2 mentions)
Holey carbon film grid, by Quantifoil (2 mentions)
Vitrobot, by Thermo Fisher Scientific (2 mentions)
Jem 1230 transmission electron, by JEOL (1 mentions)
82 protocols using ultrascan 4000 ccd camera
1
Cryo-EM Imaging of PRD1 Virus
2
Transmission Electron Microscopy of Ovarioles
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Dissected ovaries were processed for transmission electron microscopy as described previously (Liu et al., 2015 (link)). Briefly, ovarian tissue was fixed using 4% paraformaldehyde and 2% glutaraldehyde in 0.1 M sodium cacodylate (NaCac) buffer (pH 7.4). Ovarioles were then embedded within agarose. Stage 9 and 10 egg chambers were idenfied and isolated out of the agarose. The samples were post fixed in 2% osmium tetroxide in NaCac, stained with 2% uranyl acetate, dehydrated with a graded ethanol series, and embedded in EponAraldite resin. Next, thin sections were cut using a diamond knife on a Leica EM UC6 ultramicrotome (Leica Microsystems, Bannockburn, IL), collected on copper grids, and stained with uranyl acetate and lead citrate. Egg chambers were observed in a JEM 1230 transmission electron microscope (JEOL USA, Peabody, MA) at 110 kV and imaged with an UltraScan 4000 CCD camera and First Light Digital Camera Controller (Gatan, Pleasanton, CA).
3
Cryogenic HR-TEM Imaging of DF(I)NKF(I)
4
Transmission Electron Microscopy Tissue Preparation
5
Negative Staining of Protein Samples
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Protein samples were adsorbed to glow-discharged carbon-coated copper grids for 30 s prior to 2% uranyl formate staining. Micrographs were recorded using the Leginon software71 (link) on a 120 kV FEI Tecnai G2 Spirit with a Gatan Ultrascan 4000 CCD camera at 67,000 nominal magnification. The defocus ranged from 1.0 to 2.0 µm, and the pixel size was 1.6 Å.
6
Ultrastructural Analysis of Cellular Morphology
7
Quantifying Autophagic Vacuoles in Infected Cells
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Mock-infected or MCMV-infected RPE cells were treated with typsin for 5 min at 37 °C and centrifuged at 200 ×g for 5 min. After the supernatant was removed, the cell pellets were fixed in 2% glutaraldehyde in 0.1 M sodium cacodylate (NaCac) buffer, pH 7.4, postfixed in 2% osmium tetroxide in NaCac, stained en bloc with 2% uranyl acetate, dehydrated in a graded ethanol series, and embedded in Epon-Araldite resin. Thin sections were made using a diamond knife on a Leica EM UC6 ultramicrotome (Leica Microsystems, Wetzlar, Germany), collected on copper grids, and stained with uranyl acetate and lead citrate. Cells were observed under a JEM 1230 transmission electron microscope (JEOL USA, Peabody, MA) at 110 kV and imaged with an UltraScan 4000 CCD camera and First Light Digital Camera Controller (Gatan, Pleasanton, CA). Autophagic vacuoles were counted in individual cells from multiple fields and nonserial sections. Autophagic vacuoles were quantified by counting the number of autophagic vacuoles per cell.
8
Cryo-EM Structural Analysis of ChsH1-ChsH2-Ltp2
9
Cryo-electron Microscopy of Vitrified Biomolecules
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Imaging
was carried out using a field emission cryo-electron microscope
(JEOL JEM-3200FSC), operating at 200 kV. Images were taken in bright
field mode and using zero loss energy filtering (omega type) with
a slit width of 20 eV. Micrographs were recorded using a Gatan Ultrascan
4000 CCD camera. The specimen temperature was maintained at −187
°C during the imaging. Vitrified specimens were prepared using
an automated FEI Vitrobot device using Quantifoil 3.5/1 holey carbon
copper grids with a hole size of 3.5 μm. Just prior to use,
grids were plasma cleaned using a Gatan Solarus 9500 plasma cleaner
and then transferred into the environmental chamber of a FEI Vitrobot
at room temperature and 100% humidity. Thereafter, 3 μL of the
sample solution was applied on the grid and it was blotted twice for
5 s and then vitrified in a 1:1 mixture of liquid ethane and propane
at a temperature of −180 °C. The grids with vitrified
sample solution were maintained at liquid nitrogen temperature and
then cryo-transferred to the microscope.
was carried out using a field emission cryo-electron microscope
(JEOL JEM-3200FSC), operating at 200 kV. Images were taken in bright
field mode and using zero loss energy filtering (omega type) with
a slit width of 20 eV. Micrographs were recorded using a Gatan Ultrascan
4000 CCD camera. The specimen temperature was maintained at −187
°C during the imaging. Vitrified specimens were prepared using
an automated FEI Vitrobot device using Quantifoil 3.5/1 holey carbon
copper grids with a hole size of 3.5 μm. Just prior to use,
grids were plasma cleaned using a Gatan Solarus 9500 plasma cleaner
and then transferred into the environmental chamber of a FEI Vitrobot
at room temperature and 100% humidity. Thereafter, 3 μL of the
sample solution was applied on the grid and it was blotted twice for
5 s and then vitrified in a 1:1 mixture of liquid ethane and propane
at a temperature of −180 °C. The grids with vitrified
sample solution were maintained at liquid nitrogen temperature and
then cryo-transferred to the microscope.
10
Ultrastructural Analysis of CPI-613 Effects
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Approximately 1.0 × 107 cells treated with 200 μM CPI-613 or vehicle were fixed with 2% glutaraldehyde in 0.1 M sodium cacodylate (NaCAC) buffer (pH 7.4) for 45 min. The samples were post-fixed in 2% osmium tetroxide in NaCAC, stained with 2% uranyl acetate, dehydrated with a graded ethanol series and embedded in Epon-Araldite resin. Thin sections were cut with a Leica EM UC6 ultramicrotome (Leica Microsystems), collected on copper grids, and stained with uranyl acetate and lead citrate. Cells were observed in a Hitachi HT7700 transmission electron microscope and imaged with an UltraScan 4000 CCD camera and First Light Digital Camera Controller (Gatan).
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