Nontoxic nanopore electroporation for effective intracellular delivery of biological macromolecules

Significance Efficient nonviral delivery of macromolecules including mRNA, DNA plasmids, Cas9 ribonucleoproteins, and functional protein into both adherent cells and suspension cells with high cell viability is crucial for cellular manipulation, cellular imaging, and medical applications. However, the conventional delivery methods are limited to a certain range of molecules and cell types and often reduce cell viability. Here, we demonstrate effective delivery of macromolecules by nanopore electroporation (NanoEP) using a water-filter nanoporous membrane. As compared with conventional electroporation that porates the entire cell, NanoEP induces localized electroporation on a nanosized area of the cell membrane, which preserves cell viability. Additionally, our delivery method does not require specialized equipment, which allows for easy access across laboratories and medical facilities.