Atomic Force Microscopy (AFM) Imaging

The BLM samples were imaged with AFM in the PeakForce quantitative nanomechanical mapping (PF-QNM) mode using a MultiMode 8 system (Bruker) equipped with an E scanner. The system was turned on and allowed to equilibrate for at least 30 min before each experiment. The BL-AC40TS (Olympus) and RTESPA300 (Bruker) cantilevers with a spring constant of 0.1 and 40 N m^–1, and the resonance frequency of 50 and 300 kHz, respectively, were used for sample imaging in liquid and air, respectively. The AFM cantilevers were cleaned by consecutive immersing in a detergent bath, 2-propanol, and Milli-Q water for 10 min. Next, the cantilevers were ozonized in the UVC-1014 UV ozone cleaner (Nanobioanalytics, Berlin, Germany) for 10 min. The cantilevers were calibrated using the thermal tune method. The tip radius was determined by imaging the Ti roughness sample (Bruker) routinely used for tip radius determination.^69,70 The V1 grade mica disks (Ted Pella, Inc.) were mounted on metallic disks using an adhesive tape. Next, mica was cleaned in ethanol and then in Milli-Q water. After drying with an Ar stream, its top layer was piled off using an adhesive tape, resulting in a clean and atomically flat surface. The samples were immediately deposited on the freshly cleaned mica surface.

The PF-QNM in a fluid mode was used to study the morphology of BLM, BLM-AβOs, and BLM-AβOs-K162 in the PBS solution (pH = 7.4) at 21 °C. Before the imaging, the fluid cell and AFM accessories were cleaned in a detergent bath, followed by sequential rinsing with ethanol and then Milli-Q water. A 30-μL aliquot of the lipid vesicle solution (either without or with AβOs and K162) was deposited on a freshly cleaved mica substrate and then left for 45 min to form BLM on the substrate surface. Finally, the sample was rinsed with Milli-Q water filtered through a Whatman syringe filter (GE Healthcare Life Sciences) of 0.02 μm porosity and then mounted for AFM imaging.

The PF-QNM in air mode was used for monitoring Aβ aggregation in the absence and presence of K162. A freshly prepared AβMs solution (either without or with K162) was deposited on a freshly cleaved mica substrate. After 5 min of deposition, the sample was rinsed with filtered Milli-Q water, dried with a gentle stream of Ar, and subsequently mounted for AFM imaging. The imaging was performed at 21 °C.

All AFM images were processed and analyzed using Gwyddion software.⁷¹