The site-selective functionalization of mesoporous nanoparticles was achieved with a sequential co-condensation approach. Using this strategy, functional groups were completely dispersed inside the channels, concentrated in parts of the mesopores, or exclusively placed on the external surface of colloidal mesoporous silica nanoparticles depending on the time of addition of the desired functionality. This method overcomes some of the inherent disadvantages often found in selective functionalization by grafting approaches, that is, partial diffusion of reactant species inside the channels and the lack of control over the amount of incorporated organic moieties. Furthermore, the functional group density on the outer particle surface can be easily adjusted by variation of the organosilane-to-silane ratio.