Protein crosstalks as well as protein-ligand interactions play essential roles in many biological processes including signaling pathways, transcriptional regulation and numerous other metabolic reactions. In order to understand the role of such protein interactions in biological processes it is important to investigate the interaction dynamics describing the stoichiometry of the complexes, the binding free energy and their binding cooperativity as inter-molecular communication. These biochemical parameters are complementary to structural biology studies. In particular, the enthalpic and entropic components of the binding free energy directly refer to the mechanistic aspects of the binding and have been widely exploited in drug discovery research pipeline. The aim of this course is the description of practical and theoretical aspects of biophysical methods used for measuring the stoichiometry and affinity of many protein interactions. In particular, the course will focus on the application potential of the following techniques in the field of biochemistry structural biology: i) small-angle X-ray scattering; ii) isothermal titration calorimetry (ITC); iii) differential scanning fluorescence (DSF); iv) surface plasmon resonance (SPR) and v) microscale thermophoresis (MST).