Ultrasonic contrast agents have been of heightened interest in recent years. More success has been achieved by agents consisting of micro bubbles, since only a few of these agents are capable of producing very strong ultrasonic backscattered signals for the enhancement of certain tissue structures. Recent investigations also demonstrate that an analysis of the radio frequency (RF) backscattered echoes by the contrast agents may lead to quantitative means for assessing tissue perfusion. In these studies, a parameter, mean frequency shift (MFS) of the RF signal, along with integrated backscatter (IB) has received the most attention. In an effort to better understand the physical mechanisms responsible for the observed mean frequency shift, we have performed experiments on 10 dogs following injections of Albunex ® (Molecular Biosystems, Inc.) into the left atrium, coronary artery and abdominal aorta, respectively, for investigations in the heart and kidney. The integrated backscatter and mean frequency (MF) of a region of interest (ROI) were calculated from the RF signal acquired with a modified real-time ultrasonic scanner. The results show consistently that the RF signals acquired from all regions of interest are greatly affected by the presence of the contrast agent in the path between the transducer and the ROI, which can cause either an upward or a downward shift of the MF. This could not be observed by video densitometry or a measurement of the IB alone. The MFS is the result of the resonant behavior of the micro bubbles, which is related to the frequency, ambient pressure, and physical properties of the bubbles including size distribution, surface tension and concentration. On the other hand, when there is no contrast agent present in the path, a downward frequency shift is seen.