Stem/progenitor cells participate in many pathological and regenerative processes, which can be studied in vivo with molecular imaging approaches. Labeling cells with contrast agents for non-invasive imaging and tracking typically requires long exposure times or adjunctive methods, such as electroporation or transfection agents that can compromise cell viability. We have previously utilized perfluorocarbon (PFC) nanoparticles (∼200 nm) for cell tracking, but this process entailed 12 hours of incubation to achieve efficient cellular labeling. We sought to develop an approach that reduces the prolonged labeling time by enhancing PFC nanoparticle-to-cell interactions using clinical levels of ultrasound energy. Methods: Stem/progenitor (CD34 +CD133+CD31+) cells were derived from human umbilical cord mononuclear cells grown in fibronectincoated OptiCell™ cassettes. Ultrasound (US) was applied using a medical imaging system (Acuson Sequoia) and broadband (23MHz) phased array transducer, which was coupled to cells in a heated (37°C) waterbath. The transducer was advanced mechanically to expose the entire surface to calibrated levels of US energy (MI: 1.9, frequency: 2MHz). Estimated exposure time for individual cells was ∼5min. Results: Flow cytometry revealed US application greatly improved labeling when compared with cells not exposed to US (55±9% vs. 17±12% respectively, p<0.001). This approaches the labeling efficiency achieved in the 12-hr incubation method (71±4%). Under conditions of energy (ATP) depletion (20mM sodium azide, 50mM 2-deoxyglucose), augmentation of labeling with US application was prevented (3.6±0.8% vs. 4.0±1.6%, with and without US, respectively). US exposure did not compromise cell membrane integrity based on calcein dye exclusion. Cell viability was equivalent for the untreated and US-exposed cells (∼90% viable each). We conclude that ultrasound-enhanced cell labeling with PFC nanoparticles is safe and rapid, achieving sufficient labeling with only 5 minutes of exposure at clinical frequencies and power levels. The labeling process depends on active cellular transport mechanisms (i.e., ATP) and is quite distinct from typical acoustic delivery methods, which must alter or disrupt the cell membrane to achieve delivery (e.g., sonoporation, cavitation). This unique approach should facilitate investigation of regenerative therapeutics by providing a safe adjunctive method to label cells for real-time tracking.