The separation of nanoscale particles based on their differences in size is an essential technique to the nanoscience and nanotechnology community. Here, nanoparticles are successfully separated in a continuous flow by using tilted-angle standing surface acoustic waves. The acoustic field deflects nanoparticles based on volume, and the fractionation of nanoparticles is optimized by tuning the cutoff parameters. The continuous separation of nanoparticlesis demonstrated with a ≈90% recovery rate. The acoustic nanoparticle separation method is versatile, non-invasive, and simple.