The presence of surfaces influences the kinetics of amyloid-beta (A beta) peptide fibrillation. Although it has been generally recognized that the fibrillation process can be assisted or accelerated by surface chemistry, the impact of surface topography, i.e., roughness, on peptide fibrillation is relatively little understood. Here we study the role of surface roughness on surface-mediated fibrillation using polymer coatings of varying roughness as well as polymer micro particles. Using single-molecule tracking, atomic force microscopy, and the thioflavin T fluorescence technique, we show that a rough surface decelerates the two-dimensional (2D) diffusion of peptides and retards the surface-mediated fibrillation. A higher degree of roughness that presents an obstacle to peptide diffusion is found to inhibit the fibrillation process.