Fig. 2: Focal Aggregation and Uncaging of small molecules by AU-FUS.

a Small-molecule-loaded Ultrasound-Controlled carrier (UC-carrier) design. DSPC and DSPE-PEG2k form the lipid shells of the microbubbles (monolayer) and liposomes (bilayer). The lipid microbubbles (~1.5 μm mean diameter) have perfluorobutane (PFB, C₄F₁₀) gas core and have DSPE-PEG5k-Mal on the surface for conjugation with liposomes (~116 nm mean diameter; conjugated UC-carriers have ~1.7 μm mean diameter; Supplementary Fig. 3), which contained DSPE-PEG5k-SH on the surface, with a PBS core (illustration modified from Wang et al.⁶⁹). Small molecules are actively loaded into the liposomes using repeated freeze-thaw cycles, prior to conjugation (see “Methods”). b Setup for in vitro characterization of FUS sequences consists of dye-loaded UC-carriers flowing through microdialysis tubing (13 kDa cutoff, single pass), embedded in low-melt agarose. The system is confocally aligned to an inverted water-immersion 60x objective lens, and a 2.5 MHz FUS transducer. The entire setup is inside a custom-made water tank, filled with degassed Milli-Q water. c The AU-FUS sequence design. First, the aggregation sequence with peak-negative pressure P_A and duration t_A is applied. This is immediately followed by the uncaging sequence with peak-negative pressure P_U and with a fixed number of cycles (NOC) and pulse-repetition frequency (PRF) for a total duration t_U. A reperfusion period (FUS-OFF) with duration t_OFF permits reperfusion of UC-carriers. The entire sequence is repeated for the duration of sonication (N). d UC-carriers (white particles in the image) flowing through microdialysis tubing (left) with no FUS. Aggregation (blue, middle) and uncaging (green, right) sequences concentrate the UC-carriers focally and release the small molecules, respectively (see also Supplementary Movie 1). Dashed line indicates wall of tubing. Scale bar is 50 μm.