AbstractThe behaviour of new cationic amphiphilic copolymers derived from 3‐vinylpyridine was studied in aqueous medium. The formation of hydrophobic microdomains was evidenced by fluorescence spectroscopy using a molecular rotor as a fluorescent probe and their structure was visualized by cryo‐transmission electron microcospy (Cryo‐TEM). The parameters (shape and size) obtained from Cryo‐TEM were discussed in relation to the copolymers composition and to the local viscosity defined by the rotor fluorescence quantum yield.