Yes ,It's possible to measure roughness of a selected area with related softwares.I my self use Gwyddion and find it user friendly and easy. It has some thing like ruller and give you roughness of a selected region.
Yes, AFM can measure the roughness of samples. You can measure a line scan or a rectangular area scan. The scan area depends on the AFM tool: Bruker Innova, I guess, has a 100 um x 100 um scan area. For other tools, it can be larger. But we usually scan 5um x 5 um or 10 um x 10 um area.