This study evaluates the performance of subharmonic-aided pressure estimation (SHAPE) with ultrasound contrast agents using data obtained from calibration procedures. A commercial scanner of ultrasonography (GE Healthcare Logiq E20, Wuxi, China) was used to perform SHAPE on a flowing contrast agent (Sonazoid) in a phantom setup under controlled microbubble stability and flow velocity conditions. Subharmonic time-intensity curves were collected during the SHAPE calibration procedure for analysis. Subharmonic amplitude of diluted contrast agents exhibited a time-dependent decline but was not affected by velocity. SHAPE sensitivity was measured through reciprocal pressurizing and depressurizing sequences to mitigate the effect of subharmonic decline over time. A wide range of mechanical index (MI) levels within the steady growth phase of the calibration curve showed higher SHAPE sensitivity compared to the traditionally recommended “optimal” MI at the maximum slope. The approximate maximum SHAPE sensitivity was −0.04 dB/mm Hg. Subharmonic amplitude was linearly correlated with pressure at a range of MI levels (R2 > 0.9, p < 0.05) but showed significant variations (approximately 2 dB standard deviation) in the time series. The lowered sensitivity compared to previous reports, combined with the substantial variation in subharmonic amplitude, raises concerns about the accuracy and consistency of SHAPE in clinical applications.