The purpose of this study was to identify the pressure threshold for the destruction of Optison (octafluoropropane contrast agent; Amersham Health, Princeton, NJ) using a laboratory‐assembled 3.5‐MHz pulsed ultrasound system and a clinical diagnostic ultrasound scanner. A 3.5‐MHz focused transducer and a linear array with a center frequency of 6.9 MHz were positioned confocally and at 90° to each other in a tank of deionized water. Suspensions of Optison (5–8 × 10⁴ microbubbles/mL) were insonated with 2‐cycle pulses from the 3.5‐MHz transducer (peak rarefactional pressure, or P_r, from 0.0, or inactive, to 0.6 MPa) while being interrogated with fundamental B‐mode imaging pulses (mechanical index, or MI, = 0.04). Scattering received by the 3.5‐MHz transducer or the linear array was quantified as mean backscattered intensity or mean digital intensity, respectively, and fit with exponential decay functions (Ae^−kt + N, where A + N was the amplitude at time 0; N, background echogenicity; and k, decay constant). By analyzing the decay constants statistically, a pressure threshold for Optison destruction due to acoustically driven diffusion was identified. The decay constants determined from quantified 3.5‐MHz radio frequency data and B‐mode images were in good agreement. The peak rarefactional pressure threshold for Optison destruction due to acoustically driven diffusion at 3.5 MHz was 0.15 MPa (MI = 0.08). Furthermore, the rate of Optison destruction increased with increasing 3.5‐MHz exposure pressure output. Optison destruction was quantified with a laboratory‐assembled 3.5‐MHz ultrasound system and a clinical diagnostic ultrasound scanner. The pressure threshold for acoustically driven diffusion was identified, and 3 distinct mechanisms of ultrasound contrast agent destruction were observed with acoustic techniques.