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Safety of biomedical ultrasound largely depends on controlling cavitation bubbles in vivo, yet bubble nuclei in biological tissues remain unexplored compared to water. This study evaluates the effects of elastic modulus (E) and impurities on bubble nuclei available for cavitation in tissue-mimicking polyacrylamide (PA) hydrogels. A 1.5 MHz focused ultrasound transducer with f# = 0.7 was used to induce cavitation in 17.5%, 20%, and 22.5% v/v PA hydrogels using 10-ms pulses with pressures up to peak negative pressure (p−) = 35 MPa. Cavitation was monitored at 0.075 ms through high-speed photography at 40 000 fps. At p− = 29 MPa for all hydrogels, cavitation occurred at random locations within the −6 dB focal area [9.4 × 1.2 mm (p−)]. Increasing p− to 35 MPa increased bubble location consistency and caused shock scattering in the E = 282 MPa hydrogels; as the E increased to 300 MPa, bubble location consistency decreased ( p = 0.045). Adding calcium phosphate or cholesterol at 0.25% w/v or bovine serum albumin at 5% or 10% w/v in separate 17.5% PA as impurities decreased the cavitation threshold from p− = 13.2 MPa for unaltered PA to p− = 11.6 MPa, p− = 7.3 MPa, p− = 9.7 MPa, and p− = 7.5 MPa, respectively. These results suggest that both E and impurities affect the bubble nuclei available for cavitation in tissue-mimicking hydrogels.