Thermal and Acoustic Stabilization Of Volatile Phase-Change Contrast Agents Via Layer-By-Layer Assembly.

OBJECTIVE:Phase-change contrast agents (PCCAs) are perfluorocarbon nanodroplets (NDs) that have been widely studied for ultrasound imaging in vitro, pre-clinical studies, and most recently incorporated a variant of PCCAs, namely a microbubble-conjugated microdroplet emulsion, into the first clinical studies. Their properties also make them attractive candidates for a variety of diagnostic and therapeutic applications including drug-delivery, diagnosis and treatment of cancerous and inflammatory diseases, as well as tumor-growth tracking. However, control over the thermal and acoustic stability of PCCAs both in vivo and in vitro has remained a challenge for expanding the potential utility of these agents in novel clinical applications. As such, our objective was to determine the stabilizing effects of layer-by-layer assemblies and its effect on both thermal and acoustic stability. METHODS:We utilized layer-by-layer (LBL) assemblies to coat the outer PCCA membrane and characterized layering by measuring zeta potential and particle size. Stability studies were conducted by; 1) incubating the LBL-PCCAs at atmospheric pressure at 37∘C and 45∘C followed by; 2) ultrasound-mediated activation at 7.24 MHz and peak-negative pressures ranging from 0.71 - 5.48 MPa to ascertain nanodroplet activation and resultant microbubble persistence. The thermal and acoustic properties of decafluorobutane gas-condensed nanodroplets (DFB-NDs) layered with 6 and 10 layers of charge-alternating biopolymers, (LBL6NDs and LBL10NDs) respectively, were studied and compared to non-layered DFB-NDs. Half-life determinations were conducted at both 37∘C and 45∘C with acoustic droplet vaporization (ADV) measurements occurring at 23∘C. DISCUSSION:Successful application of up to 10 layers of alternating positive and negatively charged biopolymers onto the surface membrane of DFB-NDs was demonstrated. Two major claims were substantiated in this study; namely, (1) biopolymeric layering of DFB-NDs imparts a thermal stability up to an extent; and, (2) both LBL6NDs and LBL10NDs did not appear to alter particle acoustic vaporization thresholds, suggesting that the thermal stability of the particle may not necessarily be coupled with particle acoustic vaporization thresholds. CONCLUSION:Results demonstrate that the layered PCCAs had higher thermal stability, where the half-lifes of the LBLxNDs are significantly increased after incubation at 37∘C and 45∘C. Furthermore, the acoustic vaporization profiles the DFB-NDs, LBL6NDs, and LBL10NDs show that there is no statistically significant difference between the acoustic vaporization energy required to initiate acoustic droplet vaporization.