Quantification of Vaporised Targeted Nanodroplets Using High-Frame-Rate Ultrasound and Optics

Molecular targeted nanodroplets that can extravasate beyond the vascular space have great potential to improve tumor detection and characterisation. High-frame-rate ultrasound, on the other hand, is an emerging tool for imaging at a frame rate one to two orders of magnitude higher than those of existing ultrasound systems. In this study, we used high-frame-rate ultrasound combined with optics to study the acoustic response and size distribution of folate receptor (FR)-targeted versus non-targeted (NT)-nanodroplets in vitro with MDA-MB-231 breast cancer cells immediately after ultrasound activation. A flow velocity mapping technique, Stokes' theory and optical microscopy were used to estimate the size of both floating and attached vaporised nanodroplets immediately after activation. The floating vaporised nanodroplets were on average more than seven times larger than vaporised nanodroplets attached to the cells. The results also indicated that the acoustic signal of vaporised FR-targeted-nanodroplets persisted after activation, with 70% of the acoustic signals still present 1 s after activation, compared with the vaporised NT-nanodroplets, for which only 40% of the acoustic signal remained. The optical microscopic images revealed on average six times more vaporised FR-targeted-nanodroplets generated with a wider range of diameters (from 4 to 68 µm) that were still attached to the cells, compared with vaporised NT-nanodroplets (from 1 to 7 µm) with non-specific binding after activation. The mean size of attached vaporised FR-targeted-nanodroplets was on average about threefold larger than that of attached vaporised NT-nanodroplets. Taking advantage of high-frame-rate contrast-enhanced ultrasound and optical microscopy, this study offers an improved understanding of the vaporisation of the targeted nanodroplets in terms of their size and acoustic response in comparison with NT-nanodroplets. Such understanding would help in the design of optimised methodology for imaging and therapeutic applications.