AFM Imaging of Amyloid Fibrils

The end-point samples from the aggregation procedures used for the ThT assay were visualized using AFM imaging. Cleaved mica surfaces were incubated for 1 min with 10 μL of 0.5% (v/v) (3-aminopropyl)triethoxysilane (Sigma-Aldrich) in deionized water (Milli-Q). Then, the surfaces were rinsed three times with 1 mL of deionized water (Milli-Q) and dried with a gentle stream of nitrogen. For each sample, 10 μL of the solution was deposited on the positively functionalized surface. The droplet was incubated for 10 min, then rinsed with 1 mL of deionized water (Milli-Q) and dried with a gentle stream of nitrogen; all procedures were carried out at room temperature. AFM maps were acquired using JPK Nanowizard 2 system operating in tapping mode and equipped with a silicon tip (Micromasch, 2 Nm^–1) with a nominal radius of 10 nm. Image flattening was performed by SPIP (Image Metrology) software. Analysis of mean fibril heights was carried out as previously reported [48 (link)] and described in detail in Additional file 2: Supplementary Information.