This work explored the effect of lateral phase separation of phospholipids on acoustic contrast of perfluorocarbon droplets. Nanodroplets containing a volatile liquid perfluorocarbon core have been studied as ultrasound contrast agents owing to their ability to be vaporized into high-contrast microbubbles in response to high intensity focused ultrasound. These droplets are generally stabilized by a mixture of phospholipids and polymer-lipids that provide both stability and resistance to serum proteins. However, while there have been several studies on the effect of different acoustic parameters on contrast, the effect of the droplet’s stabilizing shell has not been studied as extensively. Inspired by previous studies showing lateral phase separation in microbubbles and vesicles, nanodroplets were formulated with a perfluorocarbon core and a shell composed of varying amounts of saturated (DPPC) and unsaturated (DOPC) phospholipids along with cholesterol. The stability of these mixed lipid droplets was confirmed via Nanoparticle Tracking Analysis of particle size and concentration. Because of difficulties in using fluorescence microscopy to measure phase separation on nanodroplets measuring less than 400 nm, lateral lipid phase separation was characterized by FRET analysis and TEM. The effect of the consequent shell composition on phase separation, coexistence, and nature of domain distribution in the monolayer was thus examined. It was found that, monolayer phase separation led to a distinct increase in acoustic response from the droplets on application of high intensity focused ultrasound by at least one order of magnitude. Droplets containing mixed lipid monolayers were also found to produce a significantly greater and less polydisperse yield than single-component droplets, which could be useful for scaling up preparation protocols. These monolayer-dependent properties may help not only to understand the formulation and behavior of acoustically responsive nanodroplets, they would also aid in tuning the interfacial properties of these contrast agents to obtain desired responses in different biological environments.

Lateral phase separation in superheated perfluorocarbon nanodroplet monolayers leading to enhanced ultrasound contrast imaging

251st National Meeting (2016)

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