Atomization and fountain formation is a well-known phenomenon that occurs when a focused ultrasound wave in liquid encounters an air interface. High intensity focused ultrasound (HIFU) has been shown to fractionate tissue into submicron-size fragments in a process termed boiling histotripsy, wherein the focused ultrasound wave superheats the tissue at the focus, producing a millimetre-size boiling or vapour bubble in several milliseconds. Yet the question of how this millimetre-size boiling bubble creates submicron-size tissue fragments remains. The hypothesis of this work is that tissue can behave as a liquid such that it forms a fountain and atomization within the vapour bubble produced in boiling histotripsy. We describe an experiment, in which a 2-MHz HIFU transducer (maximum in situ intensity of 24,000 W/cm2) was aligned with an air-tissue interface meant to simulate the boiling bubble. Atomization and fountain formation were observed with high-speed photography and resulted in tissue erosion. Histological examination of the atomized tissue showed whole and fragmented cells and nuclei. Air-liquid interfaces were also filmed. Our conclusion was that HIFU can fountain and atomize tissue. Although this process does not entirely mimic what was observed in liquids, it does explain many aspects of tissue fractionation in boiling histotripsy.

When focused ultrasound waves of moderate intensity in liquid encounter an air interface, a chain of drops emerges from the liquid surface to form what is known as a drop-chain fountain. Atomization, or the emission of micro-droplets, occurs when the acoustic intensity exceeds a liquid-dependent threshold. While the cavitation-wave hypothesis, which states that atomization arises from a combination of capillary-wave instabilities and cavitation bubble oscillations, is currently the most accepted theory of atomization, more data on the roles of cavitation, capillary waves, and even heat deposition or boiling would be valuable. In this paper, we experimentally test whether bubbles are a significant mechanism of atomization in drop-chain fountains. High-speed photography was used to observe the formation and atomization of drop-chain fountains composed of water and other liquids. For a range of ultrasonic frequencies and liquid sound speeds, it was found that the drop diameters approximately equalled the ultrasonic wavelengths. When water was exchanged for other liquids, it was observed that the atomization threshold increased with shear viscosity. Upon heating water, it was found that the time to commence atomization decreased with increasing temperature. Finally, water was atomized in an overpressure chamber where it was found that atomization was significantly diminished when the static pressure was increased. These results indicate that bubbles, generated by either acoustic cavitation or boiling, contribute significantly to atomization in the drop-chain fountain.

Significance High intensity focused ultrasound (HIFU) therapy is a promising, clinically adopted method of noninvasive tissue ablation used to treat both benign and malignant conditions. This work presents, to our knowledge, the first in vivo validation of a previously developed HIFU-based method that allows for noninvasive fractionation of targeted tissue into subcellular debris—boiling histotripsy—in a large animal model. While fractionating the targeted soft tissue, boiling histotripsy is shown to spare the adjacent connective tissue structures such as blood vessels. The process can be readily targeted and monitored by B-mode ultrasound. The resulting tissue debris are liquid, which provides a potential clinical benefit over thermal ablation in the treatment of tumors that exert uncomfortable pressure on surrounding tissues.

The inability to spatiotemporally guide proteins in tissues and efficiently deliver them into cells remains a key barrier to realizing their full potential in precision medicine. Here, we report ultrasound-sensitive fluoro-protein nanoemulsions which can be acoustically tracked, guided, and activated for on-demand cytosolic delivery of proteins, including antibodies, using clinically relevant diagnostic ultrasound. This advance is accessed through the discovery of a family of fluorous tags, or FTags, that transiently mask proteins to mediate their efficient dispersion into ultrasound-sensitive liquid perfluorocarbons, a phenomenon akin to dissolving an egg in liquid Teflon. We identify the biochemical basis for protein fluorous masking and confirm FTag coatings are shed during delivery, without disrupting the protein structure or function. Harnessing the ultrasound sensitivity of fluorous emulsions, real-time imaging is used to simultaneously monitor and activate FTag–protein complexes to enable controlled cytosolic antibody delivery in vitro and in vivo. These findings may advance the development of image-guided, protein-based biosensing and therapeutic modalities.