Several imaging techniques aimed at detecting ultrasound contrast agents (UCAs) echo signals, while suppressing signals coming from the surrounding tissue, have been developed. These techniques are especially relevant for blood flow, perfusion, or contrast dispersion quantification. However, despite several approaches being presented, improving the understanding of the ultrasound/UCAs interaction may support further development of imaging techniques. In this paper, the physical phenomena behind the formation of harmonic components in tissue and UCAs, respectively, are addressed as a possible way to recognize the origin of the echo signals. Simulations based on a modified Rayleigh, Plesset, Noltingk, Neppiras, and Poritsky equation and transmission and backscattering measurements of ultrasound propagating through UCAs performed with a single element transducer and a submergible hydrophone, are presented. Both numerical and in vitro results show the occurrence of a cumulative time delay between the second harmonic and fundamental component which increases with UCA concentration and propagation path length through UCAs, and that was clearly observable at frequencies ( f 0 ¿=¿2.5¿MHz) and pressure regimes (mechanical index¿=¿0.1) of interest for imaging. Most importantly, this delay is not observed in the absence of UCAs. In conclusion, the reported phenomenon represents a marker for UCAs with potential application for imaging.