The size dependent features of colloids at the nanometer scale have been issues of increasingly intensive research. In order to be able to correctly relate characteristics to certain size-populations, accurate and reliable particle sizing by dynamic light scattering (DLS) is a main prerequisite. So far, the complexity of the systems due to the presence of surfactants, proteins, and so forth in the nanoparticle suspensions has not been accounted for. In this work, practically relevant quantities of the frequently used PEO-PPO triblock copolymer surfactant Pluronic F-68 were studied for their effect on the size determination of nanoparticles by DLS. Induced changes in the tenside-content of the nanosphere suspension were monitored using a photometric assay and were correlated to the respective variances in mean particle size. These measurements showed that alterations in the range from 0.005 to 2% of Pluronic content are associated with shifts in diameter of 200 nm-particles by as much as 65 nm. The considerable changes that were found have been attributed to the surfactant-concentration-dependent fluctuations in the viscosity of the nanoparticle suspension, which affect the dimensions of colloids calculated according to the Stokes-Einstein relation.