Single molecule force spectroscopy (SMFS) is known to be one of the most powerful tool to investigate the relation between structure and function in molecules and proteins. The possibility to work in aqueous conditions at a single molecular level opens up an extraordinary perspective to investigate rare events at a molecular level of biological systems. Over the past years Atomic Force Microscopy (AFM) based on SMFS has provided us information, that is either difficult or impossible to get from any other method. In spite of its advancements, SMFS has not been applied to many molecules of biological relevance for several reasons, such as problems with the biological samples, data analysis and other technical issues. Indeed, the development and improvement of SMFS is becoming is very important to study biological molecules and proteins in their natural environment.
