A few small-molecule oxidants, most notably hydrogen peroxide, can act as messengers in signal transduction. They trigger so-called ‘thiol switches’, cysteine residues that are reversibly oxidized to transiently change the functional properties of their host proteins. The proteome-wide identification of functionally relevant ‘thiol switches’ is of significant interest. Unfortunately, prediction of redox-active cysteine residues on the basis of surface accessibility and other computational parameters appears to be of limited use. Proteomic thiol labeling approaches remain the most reliable strategy to discover new thiol switches in a hypothesis-free manner. We discuss if and how genomic knock-in strategies can help establish the physiological relevance of a ‘thiol switch’ on the organismal level. We conclude that surprisingly few attempts have been made to thoroughly verify the physiological relevance of thiol-based redox switches in mammalian model organisms.