All samples were subjected to SEM evaluation to study the topographical changes in implant surface and remaining bacterial biofilm. The implants were placed in fixative solution (4% paraformaldehyde, 2% glutaraldehyde in 0.1 M sodium cacodylate (NaCac)) buffer, pH 7.4, overnight and post-fixed in 2% osmium tetroxide in NaCac buffer, dehydrated through a graded ethanol series (25–100%), and critical-point dried using CO2 (Samdri790, Tousimis, Inc., Rockville, MD, USA). The dried implants were mounted on aluminum stubs with carbon adhesive tabs, electrically grounded with colloidal graphite, and sputter-coated for 6 min with gold-palladium (Anatech Hummer™6.2, Union City, CA, USA). The implants were imaged by a scanning electron microscope SEM (FEI, Inspect S50, Berno, Czech Republic), which was operated at 20 kV and electronic images were then captured. Four images were then recorded from each implant at the apical three threads using 1000×, 6000×, and 12,000× magnification. Representative images of each treatment were selected for presentation.
Hummer 6
Manufactured by Anatech
Sourced in United States
The Hummer 6.2 is a laboratory equipment product manufactured by Anatech. It is a high-performance instrument designed for specific scientific applications. The core function of the Hummer 6.2 is to perform a precise task within a controlled laboratory environment.
Automatically generated - may contain errors
Lab products found in correlation
Nanogold secondary antibody, by Nanoprobes (3 mentions)
Pelco tab, by Ted Pella (2 mentions)
Xl30 scanning electron microscope, by Thermo Fisher Scientific (2 mentions)
Whatman filter paper, by Cytiva (2 mentions)
Inspect s50, by Thermo Fisher Scientific (1 mentions)
Samdri 790, by Tousimis (1 mentions)
Aluminum stub, by Ted Pella (1 mentions)
Thermo noran system six, by Thermo Fisher Scientific (1 mentions)
Image pro plus software, by Media Cybernetics (1 mentions)
Model s 2260n, by Hitachi (1 mentions)
Jsm 5410lv scanning electron microscope, by JEOL (1 mentions)
Formvar coated 200 mesh copper grid, by Electron Microscopy Sciences (1 mentions)
Macrofire monochrome ccd camera, by Optronics (1 mentions)
Vega3, by TESCAN (1 mentions)
Freezone freeze dryer, by Labconco (1 mentions)
Fei xl30 scanning electron microscope, by Thermo Fisher Scientific (1 mentions)
P 2000, by Sutter Instruments (1 mentions)
Mp 285, by Sutter Instruments (1 mentions)
Aluminum sem stubs, by Ted Pella (1 mentions)
Inspect s, by Thermo Fisher Scientific (1 mentions)
24 protocols using hummer 6
1
Topographical Analysis of Implant Surfaces
2
Characterizing Particle Size and Morphology by SEM
3
Characterization of Resatorvid Particle Size and Morphology
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The particle size and surface morphology of raw resatorvid were obtained using scanning electron microscopy (SEM) (FEI, Brno, Czech Republic). The samples were sprinkled onto the double-sided adhesive carbon tabs (Ted Patella, Inc. Redding, CA, USA), which were adhered to aluminum stubs and coated with a 7 nm thin film of gold palladium alloy, using an Anatech Hummer 6.2 (Union City, CA, USA) sputtering system at 20 µA for 90 s under an argon plasma. The electron beam with an accelerating voltage of 30 kV was used at a working distance of approximately 9–12 mm. The SEM images were collected at various magnifications.
EDX was performed using ThermoNoran System Six (Thermo Scientific, Waltham, MA, USA) at an accumulation voltage of 30 keV; the spot size was increased until a dead time of 20–30 s was obtained.
EDX was performed using ThermoNoran System Six (Thermo Scientific, Waltham, MA, USA) at an accumulation voltage of 30 keV; the spot size was increased until a dead time of 20–30 s was obtained.
4
Electrospun Scaffold Ultrastructure Analysis
5
Floral Development of Acacia fimbriata
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Floral buds at several stages of development of A. fimbriata (Voucher N. Pabón-Mora 242, NY) were collected from the living collections at the Nolen greenhouses (NYBG) and at the Universidad de Antioquia (UdeA), fixed in 70% ethanol, and dissected in 90% ethanol. The samples were then dehydrated in a series of 100% ethanol, 50:50% ethanol–acetone, and 100% acetone, critical point-dried using a Samdri 790 CPD (Rockville, MD, USA), coated with gold and palladium using a Hummer 6.2 (Anatech, Springfield, VA, USA) sputter coater, and examined and photographed at 10 kV in a Jeol JSM-5410 LV scanning electron microscope.
6
Transmission Electron Microscopy of Purified Viruses
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Concentrated viral lysates (>108 viruses ml-1) were CsCl-purified (Duhaime et al., 2011 (link)) and 5 μl deposited on formvar coated 200 mesh copper grids (Electron Microscopy Sciences, Hatfield, PA, United States) that had been glow discharged for 3 min with a sputter coater (Hummer 6.2, Anatech, Union City, CA, United States). Grids were then stained with three drops of 0.02 μm-filtered 2% (w/v) uranyl acetate and for 30 s followed by three 10-s washes in ultra-pure water. All liquid was wicked away with filter paper to achieve negatively stained viral specimen. Grids were left to dry overnight in a desiccator at ambient temperature. Dry grids were visualized with a transmission electron microscope (Philips CM12, FEI, Hillsboro, OR, United States) at 80 kV accelerating voltage and 65,000–100,000 magnification. Micrographic images were collected using a Macrofire Monochrome CCD camera (Optronics, Goleta, CA, United States).
7
Exosome Characterization by SEM
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As we described previously [17 (link)], the EXO sample was fixed in 4% paraformaldehyde in 0.1M cacodylate buffer PH 7.4 overnight. Five microliters (5 µL) of suspended EXO preparation were applied to a carbon-Formvar coated 200 mesh nickel grid and allowed to stand for 30 min. The excess sample was wicked off onto Whatman filter paper. Grids were floated EXO side down onto a 20 µL drop of 1M Ammonium Chloride for 30 min to quench aldehyde groups from the fixation step. Grids were floated on drops of blocking buffer (0.4% BSA in PBS) for 2 h, then rinsed 3× with PBS (5 min each). Grids were set up as follows and allowed to incubate in blocking buffer or the primary antibody anti-CD63 (#PA5-92370) (Thermofisher Scientific, Waltham, MA, USA) for 1 h. Grids were floated on drops of 1.4 nm secondary antibody nanogold (Nanoprobes, Inc., Yaphank, NY, USA) diluted 1:1000 in blocking buffer for 1 h. Grids were rinsed 3× for 5 min each with DI. For visibility in the electron SEM, the exosome sample was postfixed in 2% osmium tetroxide in NaCac buffer and dehydrated in ethanol. Then, the sample was mounted on aluminum stubs and sputter-coated for 6 min with gold–palladium (Anatech Hummer 6.2, Union City, CA, USA). EXO were observed and imaged at 10 kV using an FEI XL30 scanning electron microscope (FEI, Hillsboro, OR, USA).
8
Characterizing Porous Cryogel Scaffolds via SEM
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Scanning electron microscopy (SEM; VEGA3; TESCAN, Brno, Czech Republic) was used to observe the pore structure on the cryogels. Combined scaffolds were frozen at −80 °C for 1 h prior to being lyophilized (FreeZone Freeze Dryer, Labconco, Kansas City, MO, USA) overnight. The samples were then mounted on an aluminum stub and sputter coated (HUMMER 6.2; Anatech, Sparks, NV, USA) for 240 s in gold at 15 mA under the pulse setting to avoid overheating. SEM was then used to obtain images at 100, 200, 500, and 1000× or all combined scaffolds.
ImageJ was used to analyze the pore diameter in the combined scaffolds. First, the line function was selected and was used to determine the scale via the scale bar in the SEM image. The unit of length was adjusted to microns for accurate measurements. Next, the selection tool was used to measure the length of a representative pore, specifically focusing on the long diameter. The measurement was recorded and the process was repeated 60 times, taking 15 measurements from each quadrant of the image. The data were saved in an Excel file and the length values were used for statistical analysis.
ImageJ was used to analyze the pore diameter in the combined scaffolds. First, the line function was selected and was used to determine the scale via the scale bar in the SEM image. The unit of length was adjusted to microns for accurate measurements. Next, the selection tool was used to measure the length of a representative pore, specifically focusing on the long diameter. The measurement was recorded and the process was repeated 60 times, taking 15 measurements from each quadrant of the image. The data were saved in an Excel file and the length values were used for statistical analysis.
9
Exosome Characterization via Electron Microscopy
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As described previously [12 (link),13 (link)], exosome samples were fixed overnight in 4% paraformaldehyde in 0.1M cacodylate buffer at PH 7.4. On a carbon–Formvar-coated 200 mesh nickel grid, five microliters (5 µL) of suspended exosome preparation was applied and allowed to stand 30 min. Whatman filter paper was used to wick off the excess sample. With the exosome side down, grids were floated on a 20 µL drop of 1M ammonium chloride for 30 min to quench aldehyde groups from fixation steps. Afterward, grids were floated on drops of blocking buffer (0.4% BSA in PBS) for 2 h and then rinsed 3 times for 5 min each with PBS. Grids were set up as follows and were allowed to incubate in blocking buffer or primary antibody (anti-TGFb, anti-IL10, or anti-CD63) for 1 h. For 1 h, grids were floated on drops of 1.4 nm nanogold secondary antibody (Nanoprobes, Inc., Yaphank. NY, USA), which was diluted at 1:1000 in blocking buffer. Grids were then rinsed 3 times for 5 min each with DI H2O. Exosome samples were postfixed in 2% osmium tetroxide in NaCac buffer and dehydrated in ethanol for visibility in electron SEM. Then, samples were mounted on aluminum stubs and sputter-coated for 6 min with gold–palladium (Anatech Hummer 6.2, Union City, CA, USA). Using an FEI XL30 scanning electron microscope (FEI, Hillboro, OR, USA), exosomes were observed and imaged at 10 kV.
10
Flexible and Stiff Probes for Mechanical Stimulation
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