Intravascular tracers in the blood circulation can provide a description of the flow field over time and space. To address the limitations of existing intravascular tracers, we have developed fluorescent nanoparticles capable of providing detailed information regarding the intravascular flow field. The nanoparticles were designed to maximize plasma half-life as well as minimize interactions with other blood components. The bioavailability of the particles in the blood circulation required nanoscale size and low surface charge density. Intravital imaging of nanoparticles in the microcirculation demonstrated that the fluorescence intensity of the nanoparticles was a major determinant of both temporal and spatial resolution of the flow field. We conclude that nanoparticles prepared with these physical and optical properties can provide an accurate description of the localized intravascular flow field.