BioAFMviewer AFM Analysis Arena A3 - an interactive software platform for automated analysis of HS-AFM imaging

High-speed atomic force microscopy (HS-AFM) experiments provide unique insights into nanoscale biological processes by directly visualizing the surface topography dynamics of biomolecules, such as proteins, over extended periods of time and generating large imaging data sets. However, the lack of computational methods for automated data analysis has thus far limited the full utilization of these data for the quantitative understanding of single-molecule dynamic. Here, we present the AFM Analysis Arena (A3) within BioAFMviewer as a software platform for automated, high-throughput quantitative analysis of AFM topographic imaging data within a user-friendly interactive graphical interface, implementing a convenient workflow from raw HS-AFM (AFM) imaging to statistical analysis. In several applications to HS-AFM imaging data, we demonstrate methods of automatized molecule detection and dynamic tracking to highlight the available capacities, aiming to transform HS-AFM measurements from observations towards an analytical tool in nanoscale biology.