Programmable ultrasonic fields enhance intracellular delivery in cell clusters

Abstract: Intracellular delivery of biomolecules remains a critical challenge in both basic cell biology and translational therapeutics. We introduce Programmable Acoustic Standing-wave Transfection (PAST), a microfluidic tool that leverages dynamically programmable ultrasonic fields to transiently permeabilize cell membranes and enhance biomolecular transport within cell clusters. By generating programmable acoustic potential landscapes, PAST drives cells through cycles of hydrodynamic and acoustic stresses that induce reversible pore formation, enabling diffusion-based delivery without chemical carriers or contrast agents. Experimental studies demonstrate controlled influx and efflux dynamics across multiple biomolecular species, with transport rates tunable via acoustic power, frequency modulation, and duty cycles. Theoretical scaling and numerical simulations reveal that membrane tension, pore energetics, and acoustic field distributions collectively govern transmembrane transport of biomolecules. Post-treatment assays confirm high cellular viability and sustained proliferation, underscoring the biocompatibility of the method. Remarkably, effective diffusivity estimates derived from model predictions closely match experimental transport timescales. Together, these findings establish PAST as a programmable, high-throughput, and non-invasive intracellular delivery platform, offering new opportunities for precision drug screening, gene editing, and mechanistic exploration of cellular membrane biophysics.