The size and surface of baricitinib encapsulated nanoparticles were characterized by Scanning Electron Microscopy (Zeiss EVO LS10; Cambridge, UK). The dried nanopartilces were suspended in one ml of deionized water and sonicated for 1 min to break the agglomeration if any. A drop of nanoparticle suspension was fixed on the carbon tape (SPI Supplies, West Chester, PA) and gold coated under vacuum (Q150R sputter coater, Quorum Technologies Ltd, East Sussex, UK) in an argon atmosphere at 20 mA for 60 s (Alshehri et al., 2017 (link)).
Q150r sputter coater
Manufactured by Quorum Technologies
Sourced in United Kingdom
The Q150R sputter coater is a laboratory equipment used for depositing thin films of materials onto substrates. It utilizes the process of sputter deposition, where atoms from a target material are ejected and then deposited onto the substrate, creating a uniform coating.
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4 protocols using q150r sputter coater
1
Characterizing Baricitinib Nanoparticle Size
2
Characterization of Pellet Size Distribution
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The average pellet diameter was measured using an EVO LS10 scanning electron microscope (Carl Zeiss; Cambridge, United Kingdom). Samples were gold-coated by Q150R sputter coater (Quorum Technologies Ltd, East Sussex, UK). The process was carried out in an argon atmosphere, at 20 mA for 1 min and involved applying 3–10 KV excitation electron energy [30 (link)]. Sieve analysis study was performed for pellet size distribution. To perform the analysis, the pellets were passed through stacks of sieves of 600, 800, 1000, 1250, 1600, 1700 and 2000 μm [31 (link)]. A minimum of three replicates was considered for each sample.
3
Scanning Electron Microscopy of Organoids
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Organoids were removed from BME, washed with excess AdDF+++, fixed for 15 min with 1% (v/v) glutaraldehyde (Sigma) in phosphate‐buffered saline (PBS) at room temperature and transferred onto 12‐mm poly‐L‐lysine‐coated coverslips (Corning).
Samples were subsequently serially dehydrated by consecutive 10‐min incubations in 2 ml of 10% (v/v), 25% (v/v) and 50% (v/v) ethanol–PBS, 75% (v/v) and 90% (v/v) ethanol–H2O, and 100% ethanol (2×), followed by 50% (v/v) ethanol–hexamethyldisilazane (HMDS) and 100% HMDS (Sigma).
Coverslips were removed from the 100% HMDS and air‐dried overnight at room temperature.
Organoids were manipulated with 0.5‐mm tungsten needles using an Olympus SZX9 light microscope and mounted onto 12‐mm specimen stubs (Agar Scientific).
Following gold coating to 1 nm using a Q150R sputter coater (Quorum Technologies) at 20 mA, samples were examined with a Phenom PRO tabletop scanning electron microscope (Phenom‐World).
Samples were subsequently serially dehydrated by consecutive 10‐min incubations in 2 ml of 10% (v/v), 25% (v/v) and 50% (v/v) ethanol–PBS, 75% (v/v) and 90% (v/v) ethanol–H2O, and 100% ethanol (2×), followed by 50% (v/v) ethanol–hexamethyldisilazane (HMDS) and 100% HMDS (Sigma).
Coverslips were removed from the 100% HMDS and air‐dried overnight at room temperature.
Organoids were manipulated with 0.5‐mm tungsten needles using an Olympus SZX9 light microscope and mounted onto 12‐mm specimen stubs (Agar Scientific).
Following gold coating to 1 nm using a Q150R sputter coater (Quorum Technologies) at 20 mA, samples were examined with a Phenom PRO tabletop scanning electron microscope (Phenom‐World).
4
Characterization of Nanoparticle Properties
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The size and zeta potential of the systems were determined by photon correlation spectroscopy (PCS) and electrophoretic laser Doppler anemometry, respectively, using a Zetaplus analyser system (Brookhaven Instruments Corporation, NY, USA). The nanoparticles mean diameter was determined after dispersion in ultrapure water and measured at 25ºC by dinamic light scattering angle of 90ºC, and the surface charge was determined by dilution of 200 μL the samples in 2 mL of a 0.1 mM KCl solution (pH 7.4). The yield of the process was calculated by gravimetry as described previously (Arbós et al., 2003) . The morphology and shape of the nanoparticles were examined and microphotographed by field emission scanning electron microscopy (FESEM) in a Zeiss Ultra Plus scanning electron microscope (Carl Zeiss SMT, Oberdochen, Germany) operating between 1 and 2 kV from 3 mm distance. Prior analysis and in order to enhance the quality of the images, particles were washed to remove the cryoprotector. For this purpose, freeze-dried nanoparticles were resuspended in ultrapure water and centrifuged at 27,000xg for 10 min. Then, the supernatants were rejected and the obtained pellets were mounted on copper grids. Finally, the pellet was shaded with a gold/palladium (Au/Pd) layer in a Q150R Sputter-Coater (Quorum Technologies, Ashford, UK).
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