To prepare samples for TEM, E. coli was cultured in TSB medium overnight and treated with 200 or 400 µM SA for 24 h at 35°C. Untreated and SA-treated bacteria were harvested by centrifugation at 415 × g and 25°C. Colonies of bacteria were fixed with 2.5% glutaraldehyde and 1% OsO4 fixative solution (pH 7.4) for 4 h at 4°C, and then rinsed with PBS. The samples were dehydrated with increasing concentrations of ethanol (30, 50, 70, 80, 90 and 100%). Following drying for 24 h, the samples were serially sectioned for TEM analysis at a magnification of ×60,000. Ultrathin sections (50 nm) were visualized using a transmission electron microscope (1200 EX; JEOL, Ltd. pan). Images were acquired using an AMT 2k CCD camera (Advanced Microscopy Techniques, Corp.) under standard conditions.
Amt 2k ccd camera
Manufactured by Advanced Microscopy Techniques
Sourced in United States
The AMT 2k CCD camera is a high-resolution digital imaging device designed for advanced microscopy applications. It features a 2048 x 2048 pixel sensor and can capture detailed images at high frame rates. The camera utilizes charge-coupled device (CCD) technology to provide reliable and precise image data.
Automatically generated - may contain errors
Lab products found in correlation
Tecnai g2 spirit biotwin, by Thermo Fisher Scientific (2 mentions)
1200ex electron microscope, by JEOL (2 mentions)
Fei tecnai g2 spirit bio twin tem, by Thermo Fisher Scientific (1 mentions)
Reichert ultracut s microtome, by Leica (1 mentions)
Aclar, by Ted Pella (1 mentions)
Tecnai g2 spirit biotwin electron microscope, by Advanced Microscopy Techniques (1 mentions)
Dulbecco phosphate buffered saline (dpbs), by Corning (1 mentions)
9 protocols using amt 2k ccd camera
1
TEM Analysis of E. coli Salicylic Acid Response
2
Ultrastructural Analysis of Platelet-Derived Microparticles
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Following shear exposure or sonication, GFP (100,000 platelets/μL, 2.5 mM CaCl2) were fixed with 2.5% formaldehyde, 2.5% glutaraldehyde in 0.1 M sodium cacodylate buffer, pH 7.4, for 30 min and then centrifuged (1500× g, 15 min) to sediment intact platelets (platelet-rich low-speed fraction). The supernatant was collected and centrifuged (20,000× g, 30 min) to obtain the PDMP-rich pellet (high-speed fraction). The platelet- and PDMP-rich pellets were stored separately in 0.2 M sodium cacodylate buffer at 4 °C until TEM imaging [73 (link)]. Ultrathin sections (60 nm) were prepared as previously described [42 (link)] and examined via JEOL 1200EX (JEOL USA, Peabody, MA, USA) or FEI TecnaiG2 Spirit BioTWIN (FEI, Hillsboro, OR, USA) transmission electron microscopes at 80 kV. Images were captured with an AMT 2k CCD camera (Advanced Microscopy Techniques Corp., Danvers, MA, USA).
3
Visualizing Staphylococcus aureus Extracellular Vesicles
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Purified S. aureus EVs were visualized by TEM as described [24 (link)]. For immunogold labeling of intracellular EVs, differentiated THP-1 cells were incubated with JE2 EVs for 30 min before the cells were washed with PBS and pelleted by centrifugation at 3000× g. The pelleted cells were fixed in 4% paraformaldehyde for 2 h at room temperature, and then placed in PBS containing 0.2 M glycine to quench free aldehyde groups. The fixed cell pellets were cryoprotected by incubating with PBS containing 2.3 M sucrose, frozen in liquid nitrogen, and sectioned on an ultramicrotome at −120 °C. The sections were transferred to formvar/carbon coated copper grids and stained with antibodies to S. aureus EVs, followed by a protein A-gold (5 nm) conjugate. The grids were counterstained and embedded by incubation with 0.3% uranyl acetate in 2% methyl cellulose. The samples were imaged on a JEOL1200EX electron microscope (JEOL, Peabody, MA, USA) equipped with an AMT 2k CCD camera (Advanced Microscopy Techniques Corp., Danvers, MA, USA).
4
Negative Staining Electron Microscopy
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Purified MVs were adsorbed onto Formvar/carbon coated copper grids and negatively stained with 1% uranyl acetate. The samples were imaged on a JEOL1200EX electron microscope (JEOL, Peabody, MA) equipped with an AMT 2k CCD camera (Advanced Microscopy Techniques Corp., Danvers, MA).
5
Transmission Electron Microscopy of EVs
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In total, 5 µl EV preparation was fixed with 5 µl 4% paraformaldehyde at 4°C for 30 min and added onto formvar-coated copper grids to settle for 30 min. Subsequently, the samples were fixed with 1% glutaraldehyde for 5 min at 25°C, then stained with 2% uranyl oxalate for 5 min at 25°C and 1.8% methyl cellulose uranyl acetate for 10 min on ice in the dark, embedded in epoxy resin and polymerized at 35°C for 12 h, 45°C for 12 h and 60°C for 24 h. Residual liquid was removed from the grid with filter paper and the samples were visualized at 120 kV using the FEI Tecnai G2 Spirit Bio TWIN TEM (FEI; Thermo Fisher Scientific, Inc.). Images were acquired using an AMT 2k CCD camera (Advanced Microscopy Techniques, Corp.).
6
Ultrastructural Analysis of T. cruzi-Infected NHDF Cells
7
Ultrastructural Analysis of Intraplantar Muscle
8
Negative Stain Transmission Electron Microscopy
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The material collected on the filters was resuspended using DPBS (Corning), and further pelleted by 100,000×g UC for 80 min at 4 °C. The material diluted in DPBS was added to a glow-discharged carbon-coated grid. The grids were washed with distilled water, stained with 0.75% uranyl formate, examined using Tecnai G2 Spirit BioTWIN microscopy (FEI, OR), and images recorded by the AMT 2k CCD camera (Advanced Microscopy Techniques, MA) at the Harvard Medical School EM Facility.
9
Ultrastructural Analysis of Bacterial Inactivation
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The EWNS exposed (45 mins at EWNS aerosol concentration of 40,000 #/cm3) and the unexposed bacterial samples of E. coli, S. enterica and L. innocua were pelleted to assess the inactivation mechanism. The pellets were fixed for 2 h at RT in 2.5% glutaraldehyde, 1.25% paraformaldehyde, and 0.03% picric acid in 0.1 M sodium cacodylate buffer (pH7.4) fixative. After washing they were post fixed with 1% osmiumtetroxide (OsO4)/1.5% potassiumferrocyanide (KFeCN6) for 2 h, washed in water 3 times and incubated in 1% uranyl acetate for 1 h followed by two washes in water and subsequent dehydration for 10 minutes each in 50%, 70%, 90%, 100% alcohol. The samples were then put in propyleneoxide for 1 h and infiltrated in a 1:1 mixture of propyleneoxide and TAAP Epon (Marivac Canada Inc. St. Laurent, CA). The samples were embedded in TAAB Epon and polymerized at 60 °C for 48 h. The solidified pelleted resins were sliced and imaged by TEM using JEOL 1200EX (JEOL, Tokyo, Japan), a conventional transmission electron microscope equipped with an AMT 2k CCD camera (Advanced Microscopy Techniques, Corp., Woburn, MA, USA).
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