Rtesp

Uncoated niosome (Nio), uncoated niosome with pentamidine (NioP), CG-Nio, CG-NioP, mixture of CG-Nio with mucin, and mixture of CG-NioP with mucin, were characterized. Particle diameter, polydispersity index (PDI), and ζ-potential were measured and analyzed in HEPES buffer by dynamic light scattering (DLS) using a Zetasizer (Nano ZS90, Malvern, UK) (n = 3 repeat measurements for each sample) [18 (link)].
In order to evaluate the size, the morphology and the homogeneity of the samples, atomic force microscopy (AFM) analysis has been performed using a Dimension Icon (Bruker Inc., Billerica, MA, USA) system. The samples have been prepared by depositing a drop of the niosomes solution after suitable dilution in HEPES on a clean monocrystalline Si (111) wafer. Images (20 μm × 20 μm) of the samples were acquired in standard tapping mode using standard Si probes (RTESP, Bruker Inc., Billerica, MA, USA) in air and at room temperature conditions. The diameter of imaged niosomes was determined by measuring the maximum height (corresponding to the center of the niosome) in respect to the plane of the substrate.

The cones from the in vivo fed ticks and from the artificially membrane fed ticks were sectioned along their lengths via cryo-microtoming. The thickness of each section was 20 (μm. The sections were then analyzed via AFM-QNM. AFM-QNM was performed with a Dimension Icon (Bruker) instrument in tapping mode. Silicon nitride probes (RTESP from Bruker) with a typical resonance frequency of 324–358 kHz, spring constant of 20–80 N/m, length of 115–135 μm, and tip radius of 8 nm were utilized for the imaging. A relative calibration method using a standard polystyrene film of 2.7 GPa (from Bruker) was utilized. The AFM images were captured at a scan rate of 1 Hz and 256 × 256 pixels of data points were collected. Images were taken at different locations (at least three) across the surface. NanoScope 5.30r2 software was used to capture the images. The height images were analyzed via NanoScope Analysis 1.5 (Bruker) image processing software.

For AFM imaging, protein-DNA binding mixtures containing 50 nM pUC18 plasmid DNA and 15 nM-30 nM SIRV2_Gp1 protein were prepared in adsorption buffer (40 mM HEPES pH 6.9, 10 mM NiCl₂) and deposited on freshly cleaved mica. After 5 min incubation, the mica surface was rinsed with deionized ultrapure water and blown dry with a gentle stream of nitrogen. Images were collected with a MultiMode (NanoScope IIIa) AFM (Bruker, Billerica, MA, USA) operated in tapping mode in air using RTESP (Bruker) AFM tips (cantilever length of 115–135 μm, width of 30–40 µm, a nominal spring constant of 20–80 N/m, and resonance frequencies in the range from 264 to 284 kHz). NanoScope Analysis v1.5 software (Bruker) was used to flatten the images, perform cross-section analyses of the complexes, and to make three-dimensional (3D) surface plots of selected complexes with a pitch of 3°.

We used an atomic force microscope (Bruker, Billerica, MA, USA) to evidence sphericity and measure the size distribution of microgels. Measurements were performed with silicon tips (RTESP, Bruker, Billerica, MA, USA). The measurement was done in the tapping mode with a frequency of 318 kHz using a controller (NanoScope V, Veeco, Plainview, TX, USA). A droplet (3 µL) of a highly diluted (ca. 0.0001 wt %) aqueous microgel solution was put on a silicon wafer (Sigert Wafer, diameter = 50.8 mm, Aachen, Germany) and dried in a dust-free environment in air.

Field Emission Scanning Electron Microscopy (FESEM) (TE-SCAN, MIRA III, Czech Republic) and Atomic Force Microscopy (AFM) (Nanowizard, JPK Ins., Germany) with cantilevers in tapping mode (RTESP, Bruker, USA) were used to investigate nano- and micro-structures of additives in binder samples. The substrate was coated with binder via the method proposed by Soenen et al^{56 (link)}. Thickness and roughness map images at a set-point z and 1–2 frames s⁻¹ were tested and then images (10 × 10 µm^{2 (link)}) evaluated using the open-source software Gwyddion⁷⁷ _. A thermal infrared camera (FLIR-T440, US) was recorded thermographic images binder samples over certain time periods.

The basal (1 cm) inflorescence stems from 7‐week‐old plants were ground under liquid nitrogen and then incubated at 70 °C in 96% (v/v) ethanol for 30 min. The pellet was successively washed with absolute ethanol, twice with 2 : 3 (v/v) chloroform: methanol, then once each with 65% (v/v), 80% (v/v) and absolute ethanol, and the remaining pellet was freeze‐dried as crude cell wall material. The crude cell wall material was suspended in ultrahigh‐purity water, placed on new mica using a pipette and dried in air overnight. The mica was glued onto a metal disc (15 mm diameter) and placed on the piezo scanner of an AFM (MultiMode VIII; Bruker). A hard tip (RTESP; Bruker) with radius of 8 nm, and spring constant of 40 N/m was used in the mechanical properties measurement. The precise spring constant was corrected by Sader method, and the deflection sensitivity was average determined by measuring a set of force–distance curves on the mica. The scan size was 10 μm × 10 μm, and 16 × 16 FD curves were collected for every measurement, and 10 different cell segments were randomly selected for mechanical measurements each sample. The Young's modulus was calculated using Hertz model of the NanoScope analysis software, and Wilcoxon test was used to test significance of average Young's modulus (He et al., 2015). Two biological replications were performed each experiment.

The topography of the protein films were investigated in air by AFM (Icon, Bruker) using Si tips on rectangular Si cantilevers (RTESP, Bruker). The proteins were adsorbed onto hydrophobic surfaces at 1 mg/mL in buffered pH conditions as previously explained (pH 6 for BSA and $\beta$ -lactoglobulin, and pH 7 for $\beta$ -casein) and room temperature. Ultra-flat gold surfaces were obtained by gold deposition onto mica [73 (link)]. After gluing the gold-coated mica on silicon wafers (gold-side down), the mica was removed with tetrahydrofuran (Sigma Aldrich). The gold substrate obtained was rinsed generously with absolute ethanol (99.8% of purity) and gently dried with N₂ gas. Immediately after, the gold surface was coated with CH₃ terminated self-assembled monolayers (SAM), by 4-h exposure to a solution of 1 mM of 1-octadecanethiol (Sigma Aldrich) in absolute ethanol. Finally, the substrates were rinsed again with absolute ethanol and dried with N₂ gas. We obtained a hydrophobic surface for the protein adsorption at the same conditions that in the QCM experiments (the amount of adsorbed protein over the PS surface was identical that over the gold-CH₃ surface).