Investigation of Optimized Treatment Conditions for Acoustic-Transfection Technique for Intracellular Delivery of Macromolecules.

Manipulation of cellular functions and structures by introduction of genetic materials inside cells has been one of the most prominent research areas in biomedicine. High-frequency ultrasound acoustic-transfection has recently been developed and confirmed by intracellular delivery of small molecules into HeLa cells at the single-cell level with high cell viability. After we proved the concept underlying the acoustic-transfection technique, treatment conditions for different human cancer cell lines have been intensively investigated to further develop acoustic-transfection as a versatile and adaptable transfection method by satisfying the requirements of high-delivery efficiency and cell membrane permeability with minimal membrane disruption. To determine optimal treatment conditions for different cell lines, we developed a quantitative intracellular delivery score based on delivery efficiency, cell membrane permeability and cell viability after 4 and 20 h of treatment. The intracellular delivery of macromolecules and the simultaneous intracellular delivery of two molecules under optimal treatment conditions were successfully achieved. We found that DNA plasmid was delivered by acoustic-transfection technique into epiblast stem cells, which expressed transient mCherry fluorescence.