Dependence of sonoporation efficiency on microbubble size: An in vitro monodisperse microbubble study

Sonoporation is the process where intracellular drug delivery is facilitated by ultrasound-driven microbubble oscillations. Several mechanisms have been proposed to relate microbubble dynamics to sonoporation including shear and normal stress. The present work aims to gain insight into the role of microbubble size on sonoporation by varying the microbubble size of monodisperse microbubble suspensions. Sonoporation experiments were performed in vitro on cell monolayers at an ultrasound frequency of 1 MHz at varying acoustic pressures (250–750 kPa) and pulse length (10, 100, 1000 cycles). Sonoporation efficiency was quantified using flow cytometry by measuring the FITC-dextran (4 kDa and 2 MDa) fluorescence intensity in 10,000 cells per experiment. We demonstrate that the bubble oscillation amplitude is nearly independent of the equilibrium bubble radius at acoustic pressure amplitudes that induce sonoporation (≥500 kPa). However, we show that sonoporation efficiency is strongly dependent on the equilibrium bubble size and most efficiently induced by off-resonance 4.7-μm radius bubbles. These 4.7-μm bubbles are an order of magnitude more efficient than the polydisperse bubbles. We discuss that for our system shear stress is highly unlikely the mechanism of action, while we show that sonoporation efficiency correlates well with an estimate of the bubble-induced normal stress.