Imaging collagen II using atomic force microscopy (AFM)

Collagen II is a fibrous protein that assembles from basic tropocollagen subunits to form extracellular supramolecular fiber networks within cartilage tissue. Tropocollagen subunits of ~300 nm in length self-assemble first into pentameric uniform microfibrils, which fuse into bigger collagen fibrils that can range from 10 nm to 500 nm in diameter. The collagen fibrils display a characteristic 67-nm repeat because of the staggering of individual collagen molecules with respect to each other. This protocol demonstrates how to prepare collagen protein samples for analysis by atomic force microscopy (AFM). It also describes the steps for generating AFM images of collagen samples during and after manipulation to analyze collagen self-assembly.