Fluorescence correlation spectroscopy (FCS) was applied to directly monitor the hydrophobic collapse of pH-responsive hairy nanoparticles at the individual particle level. To this end, fluorescent nanoparticles (hydrodynamic radius 20 nm) with polystyrene core and poly(N,N-diethylaminoethyl methacrylate) (PDEA) shell were prepared and used as a model system. Dynamic light scattering and turbidity measurements showed that the hydrophobic collapse of the hairs at high pH values is associated with strong interparticle aggregation that hinders determination of individual particles size. However, at the ultralow concentrations assessable by FCS (less than one particle per femtoliter) the aggregation was prevented. Thus, the pH-induced change in the particles size caused by the swelling or the collapse of the PDEA hairs was systematically measured and compared with that of individual freely diffusing PDEA chains under similar conditions.