Kratos axis ultra dld spectrometer

Subsamples (a few mg) were retained and air-dried. Surface chemical composition and oxidation state analyses were done by XPS via published methods (Omajali et al., 2017 (link)) using a Kratos Axis Ultra DLD spectrometer (Kratos Analytical). The samples were illuminated using an Al Kα x-ray source and the photoelectrons were collected using a hemispherical electron analyzer. Survey spectra were recorded using a pass energy of 160 eV, with the pass energy reduced to 20 eV for acquisition of the core level spectra (resolution approx. 0.4 eV). The samples were insulating, therefore a charge neutralizer was used to prevent surface charging with a low energy electron beam directed onto the sample during XPS data acquisition. Measurements were made at room temperature and at a take-off angle of 90°, to probe a depth of approx. 5–10 nm to examine bio-NPs bound to the outermost cell surfaces. Generated data were converted into VAMAS format and analyzed using the CasaXPS package (Fairley, 2013 ) employing Shirley backgrounds, mixed Gaussian-Lorentzian (Voigt) lineshapes and asymmetry parameters where appropriate. All binding energies were calibrated to the C 1s peak originating from C-H or C-C groups at 284.8 eV.

XPS spectra were recorded on a Kratos Axis Ultra DLD spectrometer (Kratos Analytical, Manchester UK). Survey spectra and Carbon 1s (C 1s) elemental spectra were collected from 300 x 700 μm areas on the parylene and SiO₂ regions of each sample. Survey spectra were collected with a pass energy of 160 eV and dwell time of 138 ms from five 180 s sweeps. Elemental spectra were collected with a pass energy of 20 eV and a dwell time of 260 ms from fifteen 60 seconds sweeps. The vacuum chamber pressure was kept below 2 x 10⁻⁹ Torr. Samples were illuminated with monochromated Aluminium K_α X-rays at 1486.69 eV and analysed with charge neutralisation.

X-ray photoelectron spectroscopy (XPS) analyses were carried out on a Kratos Axis Ultra^DLD spectrometer (Kratos Analytical Ltd., Manchester, UK), using monochromatized Al Kα radiation (hν = 1486.6 eV), with a selected X-ray power of 150 W. High resolution spectra were collected under the CAE (constant analyser energy) mode, with a pass energy of 20 eV. Fresh and thermally aged samples, without any further treatment, were compressed into self-supported pellets and then mounted on the sample holder, by means of a double-sided adhesive conducting carbon tape. The coaxial charge neutralization system developed by Kratos was employed to compensate surface charging effects. The binding energy (BE) scale was calibrated with respect to the C 1s signal, coming from adventitious carbon contamination, and set at 284.8 eV, according to the literature [52 (link)]. CasaXPS software (version 2.3.19rev1.1m, Casa Software Ltd., Devon, UK) was used for spectra processing.

The XPS analyses were performed with a Kratos Axis UltraDLD spectrometer (Kratos Analytical Ltd., Manchester, UK) using a monochromated Al X-ray source (1486.7 eV) operated at 150 W and pressure in the main chamber in the 10⁻⁹ torr range. The information collected was related to approximately the top 10 nm of the sample surface. Survey scans were performed with a 1.0 eV step size and a 160 eV analyzer pass energy, while the high-resolution scans were recorded at a 20 eV pass energy and a 0.1 eV step size. The spectra were processed using CasaXPS software (Version 2.3.20, Casa Software Ltd., Terrace Teignmouth, UK) and calibrated by setting the main C1s peak to 284.8 eV. The atomic surface concentration (atom%) was determined from the survey spectra and represents the average of the measurements taken at two different spots on each sample.

XPS was used to characterize the chemical composition of the various spin coated 25PS/75PMMA demixed surfaces, with and without 50 μm PS microspheres. XPS was carried out using a Kratos Axis Ultra DLD Spectrometer (Kratos Analytical Ltd., Manchester, UK) operating with aluminum Kα X-rays at an incident energy of 1486.6 eV. Wide energy survey scans (WESS) were acquired at a pass energy of 160 eV followed by high resolution spectra for the carbon C1s and oxygen O1s regions, respectively, at a pass energy of 20 eV. Given the insulating nature of the polymer demixed samples, in situ charge neutralization was applied via a low energy electron gun operating with a filament current of 1.95 A and a charge balance setting of 3.3 V, working in tandem with a magnetic immersion lens. Correction for any residual charging effects was made by setting the main component of the C1s peak to 284.6 eV, the value associated with adventitious carbon.

The surface chemical make-up at the different stages of modification was examined by XPS using a Kratos AXIS Ultra DLD spectrometer (Kratos Analytical Ltd., Manchester, UK) with an Al Kα X-ray source (1486.7 eV) [49 (link)]. C1s (C–C bond) peak at 284.6 eV was applied as a reference to compute all other binding energies. SEM (JSM-6701F-JEOL, Tokyo, Japan) was used to observe the substrate surfaces. They were first coated with gold by sputter-coating at 10 mA for 50 s. Images were acquired at 5 different regions per substrate at a voltage of 12 kV.
Anti-E2 antibody was used to visualize the bound-E2 on substrates. These substrates were incubated overnight in 3% bovine serum albumin (BSA) at room temperature (RT), before immersing in rabbit anti-E2 antibody for 3 h at RT. Substrates were washed in PBS thrice and incubated with anti-rabbit IgG–FITC for 30 min in dark at RT. They were then washed again in PBS. Finally, these substrates were ready to be observed under confocal laser scanning microscope (CLSM, Olympus FV1000, Tokyo, Japan). The amount of immobilized E2 was indirectly quantified by calculating the unbound E2 washed out in water. Unbound E2 was quantified using ELISA kit for E2 (DRG International, Inc., Springfield, NJ, USA) as per manufacturer’s instructions.