We demonstrate nanoparticle size measurement using time-resolved fluorescence anisotropy decay in relation to establishing a nanometrology standard. The rotational correlation time equivalent to the isotropic Brownian rotation of a fluorescent 6-methoxyquinolinium dye attached to amorphous silica nanoparticles was determined in three different LUDOX 2 colloids from the complex fluorescence anisotropy decay observed. Once competing depolarization and nanoparticle aggregation had been taken into account, good agreement was found of 4.0 ± 0.4 nm, 6.4 ± 0.5 nm and 11.0 ± 1.6 nm corresponding to the manufacturer's reported particle radii of 3.5 nm, 6 nm and 11 nm, for LUDOX SM30, AM30 and AS40 respectively. We describe the measurement science required for acquisition and interpretation of fluorescence anisotropy decay data in order to determine nanoparticle size while highlighting the limitations and useful range of measurement.