Membranotropic peptides is a class of peptides that exert their biological action at the level of cell membranes. Understanding how they interact with their different membrane binding partners (lipids, proteins, and/or glycoconjugates) is important to decipher their mechanism of action. Affinity photolabeling is a powerful method to study noncovalent interactions and provide a submolecular picture of the contacts between two interacting partners. In this review, we give a panorama of photolabeling-based studies of the interactions between membranotropic peptides and membranes using either photoreactive lipids or peptides.