Focusing and enriching submicrometer and nanometer scale objects is of great importance for many applications in biology, chemistry, engineering, and medicine. Here, we present an acoustofluidic chip that can generate single vortex acoustic streaming inside a glass capillary through using low-power acoustic waves (only 5 V is required). The single vortex acoustic streaming that is generated, in conjunction with the acoustic radiation force, is able to enrich submicrometer- and nanometer-sized particles in a small volume. Numerical simulations were used to elucidate the mechanism of the single vortex formation and were verified experimentally, demonstrating the focusing of silica and polystyrene particles ranging in diameter from 80 to 500 nm. Moreover, the acoustofluidic chip was used to conduct an immunoassay in which nanoparticles that captured fluorescently labeled biomarkers were concentrated to enhance the emitted signal. With its advantages in simplicity, functionality, and power consumption, the acoustofluidic chip we present here is promising for many point-of-care applications.