To prepare the substrates with different stiffnesses, 3 g of gelatin (type A, Sigma-Aldrich, USA) was dissolved in 10 ml of ultrapure water at 50°C under stirring to obtain a 30 weight % (wt %) gelatin aqueous solution. Microbial transglutaminase (0.4 g; mTG, Pangbo, China) was dissolved in 10 ml of ultrapure water at room temperature to obtain a 4 wt % aqueous solution. gelatin and mTG aqueous solutions were filtered and then mixed with different concentration, respectively (gelatin aqueous solution: 10, 15, 10, 10, 15, and 20%; mTG aqueous solution: 0.5, 0.5, 1.5, 2, 2, and 2%) in petri dishes at 37°C for 3 hours to obtain hydrogels with gradient stiffness of 5, 15, 25, 45, 65, and 125 kPa, respectively. The stiffness was measured by an atomic force microscope (Agilent Technologies, USA). After soaking in phosphate-buffered saline for 24 hours, 3T3 cells were seeded onto gelatin hydrogels at a concentration of 1 × 106 cells/ml.
Atomic force microscope
Manufactured by Agilent Technologies
Sourced in United States
The Atomic Force Microscope (AFM) is a high-resolution scanning probe microscope that allows for the observation and measurement of surface topography at the nanoscale level. It operates by scanning a sharp probe across a sample surface, detecting the interactions between the probe and the sample to generate a detailed 3D image of the surface.
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Lab products found in correlation
Escalab 250, by Thermo Fisher Scientific (3 mentions)
Escalab 250 spectrometer, by Thermo Fisher Scientific (2 mentions)
Gelatin, by Merck Group (1 mentions)
A1r plus, by Nikon (1 mentions)
Jsm 6390lv, by JEOL (1 mentions)
Invia reflex, by Renishaw (1 mentions)
Quartz cell, by Agilent Technologies (1 mentions)
Jem 1200 exii, by JEOL (1 mentions)
Spectrum 1000, by PerkinElmer (1 mentions)
Helios nanolab 660, by Thermo Fisher Scientific (1 mentions)
Cary e 5000, by Agilent Technologies (1 mentions)
Kbr pellets, by PerkinElmer (1 mentions)
Omcl ac160ts, by Olympus (1 mentions)
Uv 2600, by Shimadzu (1 mentions)
Nanoscope 5 system, by Bruker (1 mentions)
8 protocols using atomic force microscope
1
Gelatin Hydrogel Substrates with Gradient Stiffness
2
Characterizing Silver Nanoparticle Morphology
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The size and morphology of the synthesized silver nanoparticles was determined by using an Atomic Force Microscope (Agilent Technologies 5500, USA) Agilent 5500 operated in tapping mode.
3
Morphological Characterization of Hydrogel Sheets
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The cross-sectional morphologies of the as-prepared hydrogel sheet were observed by SEM (JSM-6390LV, JEOL) operated at 15.0 kV. The sample for SEM imaging was prepared by fracturing the freeze-dried hydrogel in liquid nitrogen and gold sputtering on the fracture surface. To confirm the gradient structure, the cross-sectional morphologies of the samples dyed by the dilute solution of sodium fluorescein (~10−6 M) and rhodamine 6G (~10−6 M) were respectively imaged by using CLSM (Nikon A1R plus, Tokyo, Japan) with an exciting wavelength of 488 nm in the single-channel mode. Raman scattering measurements were performed using a Raman system (inVia-reflex, Renishaw) with confocal microscopy under an excitation light of 514.5 nm. Atomic force microscopy (AFM) images were taken by an atomic force microscope (Agilent Technologies Inc.) operating in tapping mode using silicon cantilevers with ~300-kHz resonance frequency. The surface chemical constitutions of the hydrogel sheets were characterized by XPS (Thermo Fisher Scientific, ESCALAB 250).
4
Comprehensive Nanomaterial Characterization Techniques
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Transmission electron microscope (TEM) and atomic fluorescence microscopy (AFM) images were taken on a JEOL JEM 2100 TEM at an accelerating voltage of 200 kV and Agilent atomic force microscope, respectively. UV-visible absorption was carried out using an “Evolution” spectrophotometer. Fluorescence spectroscopy was performed on an LS 55 PerkinElmer spectrophotometer. The Fourier transform infrared spectral (FTIR) investigation was recorded on a 8900 Shimadzu HYPER. Experiments were performed by preparing KBr pellets in the frequency range of 400–4000 cm−1. X-ray photoelectron spectroscopy (XPS) analysis was performed by an ESCALAB 250 spectrometer (Thermo-VG Scientific Co., U.S.A.) with an ultrahigh vacuum generator. Powder X-ray diffraction (XRD) was analyzed by PANalytical-EMPYREAN.
5
Characterization of Cu@Pt Nanoparticles
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In this work, the UV–VIS absorption spectra of the synthesized Cu@Pt nanoparticles were obtained from a spectrophotometer Cary E 5000 in the range of 300–800 nm using a quartz cell with 10 mm of optical path length (Agilent, USA). FTIR spectra of the samples were measured using Perkin-Elmer Spectrum 1000, in attenuated total reflection mode, and using the spectral range of 4000–380 cm−1. The study also used one instrument in the diffuse reflectance mode at the resolution of 4 cm−1 in KBr pellets (Perkin Elmer, USA). The obtained Cu@Pt nanoparticles were characterized using an Atomic Force Microscope (Agilent, USA). The size and morphology of the synthesized Cu@Pt nanoparticles were characterized using a Transmission Electron Microscope JEOL JEM 1200 EXII, operating at 200 kV. Moreover, we used a Scanning Electron Microscope (HR SEM) Helios NanoLab 660 (FEI). SEM imaging was performed in the immersion mode (Thermo Fisher Scientific, USA).
6
Morphological Characterization of Mucus Treated with Surfactants
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Morphology study of original mucus and mucus treated by surfactants was performed by macroscopic observation, optical microscope and Atomic force microscope (AFM). In order to better distinguish the difference of mucus samples for macroscopic observation, native mucus and mucus treated by surfactants were centrifuged at 2000 r/min for 10 min. Optical microscope observation was performed by BI-2000 Image Analysis System (Chengdu Techman Software Co., Ltd) at X40 objective. Atomic force microscope (Agilent Technologies, USA) was used to study the morphology of mucus at micrometer scale. The samples were added to a clean mica plate and dried at room temperature. Then the samples were tested by tapping mode.
7
Atomic Force Microscopy of Sample Morphology
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An atomic force microscope (Agilent Technologies, San José, CA, USA) equipped with PicoView 1.14.3 control software was used to reveal the morphology and size of the studied objects. A small amount of the sample suspension was dropped onto a piece of mica (PELCO Mica Sheets Grade V5, 15 × 15 mm2), allowed to adsorb on the mica substrate for 5–10 min, and then rinsed with ultrapure water and allowed to dry under ambient conditions. Topographic images were obtained in tapping mode using standard silicon cantilevers (Olympus, model OMCL-AC 160TS, Olympus Corp., Tokyo, Japan) with a resonant frequency of 300 kHz and a spring constant of 26 N/m. All measurements were carried out in the air, at ambient temperature and relative humidity between 30 and 40%. The freely available software Gwyddion helped with image editing (http://gwyddion.net/ , accessed on 2 February 2020).
8
Characterization of I-Fe3O4-NPs and I-Fe3O4-NBC
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The synthesized I-Fe3O4-NPs and I-Fe3O4-NBC were characterized using advanced analytical techniques, i.e., UV–visible, FT-IR, SEM, and XRD. Morphological characters of I-Fe3O4-NPs and I-Fe3O4-NBC were analyzed by scanning electron microscope (SEM) model JSM-6380 (JEOL Electronics Company, Japan). Similarly, the crystal structure was studied by X-ray diffraction (D-8 of Bruker). The spectrophotometric study for the confirmation of I-Fe3O4-NPs was conducted by a double-beam UV–visible spectrophotometer (UV-2600, Shimadzu, Japan). FT-IR spectra of I-Fe3O4-NPs nanoparticles and their composites in the range of 400–4000 cm−1 were obtained by FT-IR spectrophotometer Thermo Electron Scientific (Madison, WI, USA) with a KBr pellet. The size and shape of I-Fe3O4-NPs and I-Fe3O4-NBC was confirmed by atomic force microscope (Agilent, Santa Clara, CA, USA). AFM imaging was performed on the NanoScope V system (Bruker Ltd, Germany). Dynamic light scattering (DLS) and zeta potential measurements were taken for particle size and adsorption activity of adsorbent using the laser scattering particle size distribution analyzer (Horiba Scientific, Kyoto, Japan) and the zeta potential analyzer (ELSZ-2000), respectively. Furthermore, a salt addition method was used to determine the point of zero charges (isoelectric point) of the adsorbent, as reported elsewhere [53 (link)].
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