*4.4. Implications of the Study*

The addition of cholesterol to the indirect DSPC-based microbubble coating increased the variability in ligand distribution, acoustic response, polydispersity, and buckle formation. These effects can be explained by the altered lipid phase distribution as described above and imply that the indirect DSPC-cholesterol microbubbles are less stable than those without cholesterol. Because the indirect DSPC-cholesterol microbubbles had heterogeneities in the form of buckles, they could not be regarded as microbubbles with a uniform lipid distribution when comparing their acoustic behavior to that of the microbubbles with heterogeneous lipid phase distribution, i.e., the direct and indirect DSPC microbubbles. Thus, reduced stability of the microbubble coating is expected when the components are all miscible and in the same LE phase, which will increase the heterogeneity of the microbubble population and thereby increase the variability of the acoustical response. Therefore, a different approach will be required to achieve a more uniform microbubble response to ultrasound, possibly by tailoring the LC phase domains, as our results suggest that differences in LC domain size can predict the relative oscillation amplitude.
