Tissue characterization based on the backscatter coefficient (BSC) can be degraded by acoustic nonlinearity. Often, this degradation is due to the method used for obtaining a reference spectrum, i.e., using a planar reference in water compared to a reference phantom approach resulted in more degradation. We hypothesize that an in situ calibration approach can improve BSC estimates in the nonlinear regime compared to using the reference phantom approach. The in situ calibration target provides a reference within the medium being interrogated and, therefore, nonlinear effects would already be contained in the in situ reference signal. Simulations and experiments in phantoms and in vivo were performed. A 2 mm diameter titanium bead was embedded in the interrogated media. An L9-4/38 probe (BK Ultrasound, Peabody, MA) and an analysis bandwidth from 4.5 to 7.4 MHz were used in experiments. Radiofrequency data from the sample, bead, and reference phantoms were acquired at a quasi-linear baseline power level and at further increments of output power. Better agreement between the BSC obtained at low power compared to high power was observed for the in situ calibration compared to the reference phantom approach.

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