Ultrasonic contrast agents are used to assist perfusion assessment based on evaluation of the time intensity curve (TIC). Previous results showed that such techniques are susceptible to the shadowing effect. To overcome this problem, a new TIC based technique using time-intensity relationships from both the inflow and the outflow of a perfused area is proposed. The technique is called the input-output TIC technique (IOTIC). It contrasts with conventional TIC techniques in that conventional techniques measure the time intensities directly from a perfused area. With both the inflow and the outflow time intensities, the microbubble concentration within the perfused area can be derived based on the fact that the difference between the number of microbubbles at the outflow and the inflow equals the time derivative of the number of microbubbles inside the perfused area. In this study, efficacy of the IOTIC technique is experimentally tested and compared with conventional techniques. Results show that the shadowing effect significantly affected flow estimation with conventional techniques. The IOTIC technique eliminates the shadowing effect and provides a good correlation between the actual flow rate and measured flow-related parameters, thus making quantitative estimation of perfusion feasible. Potential clinical applications of the IOTIC technique were also explored. One example is brain perfusion assessment where the poor acoustic access of the brain tissue may be avoided with the IOTIC technique.