Nanoscope analysis 2

A 5 µL volume of the freshly prepared NFPh emulsion, prepared as described in Section 4.1, was dispensed onto freshly cleaved mica. Mica surface (0.15 mm thick, sized 15 × 15 mm, TipsNano, Zelenograd, Russia) was used as a substrate for non-covalent adsorption.
The emulsion droplet was incubated on the mica substrate surface for 10 min. Then, the substrate was washed with 1 mL of deionized water, which was obtained using a Simplicity UV system (Millipore, Molsheim, France). The washed substrate was dried in air and subjected to AFM scanning.
The AFM images of nanosized particles were obtained with a Dimension atomic force microscope equipped with an Icon^™ scanner (Bruker, Billerica, MA, USA). The instrument is a part of the Avogadro unique research facility (http://avo.ibmc.msk.ru/ (accessed on 1 October 2023)). Scanning was carried out in the tapping mode. A short cantilever holder was used; the measurements were conducted in air. The images were recorded using the NanoScope 9.4 software (Bruker, Billerica, MA, USA). AFM images were processed using the standard NanoScope Analysis 2.0 software (Bruker, Billerica, MA, USA), Gwyddion 2.62, and Femtoscan Online software 4.8 (LLC Scientific and Production Enterprise “Center for Advanced Technologies”, Moscow, Russia; www.nanoscopy.net/en/Femtoscan-V.shtm (accessed on 6 May 2021)).

A total of 100 μL of β
₂M fibrils prepared in 10 mM NaH
₂PO
₄-H
₃PO
₄ (pH 2.5) were diluted into H
₂O, giving a final volume of 0.50 mL. Ten microliters of β
₂M fibril samples (~17 μM) were then incubated on a freshly cleaved mica surface for 5 min, followed by rinsing three times with 10 μL of pure water to remove the unbound fibrils and drying at room temperature. The fibrils on the mica surface were probed in air by the Dimension icon scanning probe microscope (Bruker, Santa Barbara, USA) with ScanAsyst mode. The measurements were realized by using SCANASYST-AIR probe with a spring constant of 0.4 N/m and a resonance frequency of 70 kHz (Bruker). A fixed resolution (256×256 data points) of the AFM images was acquired with a scan rate at 1 Hz and analyzed by using NanoScope Analysis 2.0 software (Bruker).

All of the anhydrous solvents and chemicals were used as received from the commercial suppliers, unless otherwise indicated. Styrene (≥99%) (Lot: STBJ7184) and benzoyl peroxide (75%) (Lot: MKCG5941) were purchased from Aldrich (Poznan, Poland). ¹H and ¹³C NMR spectra were recorded on Bruker (Billerica, MA, USA), AV-400 instrument (400 MHz). Chemical shifts (δ) were reported in parts per million (ppm). SEM images were obtained on a JEOL JSM-7400F scanning electron microscope (Akishima, Tokyo, Japan). AFM topographic images were obtained using a Nanoscope 9.7 Dimension ICON atomic force microscope (Bruker, Billerica, MA, USA) and the results were analysed using NanoScope Analysis 2.0 software (Bruker, Camarillo, CA, USA).
For antimicrobial assessments, tryptic soy broth (TSB) and Mueller Hinton broth (MHB) were purchased from Oxoid Ltd. (Basingstoke, UK), while yeast mold broth (YMB) was purchased from BD (Singapore). The broth solutions were prepared according to the manufacturer’s instructions. Gram-negative bacteria Escherichia coli (ATCC 8739), gram-positive Staphylococcus aureus (ATCC 6538), Gram-negative Pseudomonas aeruginosa (ATCC 9027), and fungi Candida albicans (ATCC 10231) were purchased from ATCC (Manassas, VA, USA) and re-cultured according to the suggested protocols.

A Bruker Dimension Icon Atomic Force Microscopy (AFM) system operating in tapping mode with TESPA-V2 tips was used to scan the nanocoils. A 50 µm wide field of view (FOV) was surveyed at a scan rate of 0.1 Hz to reduce tip and sample wear and improve image quality. The scans were imported into Bruker Nanoscope Analysis 2.0, sectioned, and exported as XZ plane height maps for further processing using Python. Turn width and spacing were determined using a partition threshold of 40 nm above the previous gap minimum. Mean turn width and spacing for all 14 turns and 13 gaps were plotted for comparison.

The thickness of the Mo₂CT_z MXene flakes were measured through imaging using an atomic force microscope (AFM, Dimension Icon; Bruker Corp., Billerica, MA, USA) and analysed with the supplied software (NanoScope Analysis 2.0; Bruker Corp., Billerica, MA, USA).

The fluorescence images were analyzed using ImageJ and AFM images were analyzed with Nanoscope Analysis 2.0 (Bruker Nanoscope Analysis 2.0 Santa Barbara, CA, USA).