Ultrastructural Analysis of aSyn Fibrils

To examine the ultrastructure of aSyn fibrils, negative staining electron microscope images were taken using Formvar/carbon-coated 200-mesh copper grids (Electron microscopy Sciences, Switzerland) as previously described^{94 (link)}. Activated grids were loaded with 5 µl of sample for 30 s, washed three times with ultrapure water, and then negatively stained with 1% uranyl acetate for 1 min. Excess liquid was removed, and grids were allowed to air dry. Imaging was carried out on a Tecnai Spirit BioTWIN electron microscope operating at 80 kV acceleration voltage and equipped with a digital camera (FEI Eagle, FEI). A total of 3–5 images for each sample were chosen and the length of fibrils quantified (average 50–100 nm) using the Image J software (U.S. National Institutes of Health, MD, USA; RRID:SCR_001935).