Expert researchers describe the key methods for detecting and analyzing proteins and protein motifs involved in targeting interactions, mostly from the perspective of intracellular signaling. The techniques include those designed to reveal the existence of targeting interactions and characterize their physico-chemical properties, to identify and isolate interacting proteins on the basis of the ‘bait-prey’ principle, to target domains, and to modify lipids. The methods are richly detailed for use by those researchers interested in defining and characterizing the protein–protein interactions that underlie targeting mechanisms in all areas of molecular cell biology.