Size-exclusion chromatography (SEC), coupled with differential viscometry detection (SEC/DV), is applied to the dilute suspension characterization of solvent-swollen cross-linked polymeric nanoparticles (PNPs). Cross-linked, unimolecular polymeric nanoparticles in the 5-50 nm weight-average diameter (d(w)) range were prepared by batch and semibatch microemulsion polymerization techniques and isolated. SEC and SEC/DV characterization techniques yield, based on the principle of universal calibration, a wealth of information regarding the structural attributes of PNPs, including apparent and absolute molecular weight distributions, apparent and absolute molecular weight averages, peak and weight-average particle diameters, particle size distributions in both the solvent-swollen and solvent-free states, particle conformation (shape), and an estimate of the volumetric swell factor. These structural parameters are critical to understanding PNP performance, and all are obtained in a single rapid chromatographic experiment, when conducted under conditions where universal calibration applies. Particle sizes determined under such conditions are in excellent agreement with those obtained by dynamic light scattering, transmission electron microscopy, hydrodynamic chromatography, and SEC/static light scattering (SEC/SLS). In addition, Mark-Houwink exponents of approximately zero were found across the molecular weight and size distribution of many of these tightly cross-linked PNPs, which is consistent with a spherical particle conformation in these dilute suspensions. The SEC/DV methods are especially valuable to characterize the diameter, volume swell factor, and suspension conformation of small (4-5 nm d(w)) PNPs.