Robust three-dimensional nanotube-in-micropillar array electrodes to facilitate size independent electroporation in blood cell therapy

Blood is an attractive carrier for plasmid and RNA-based medicine in cell therapy. Electroporation serves as a favorable delivery tool for simple operation, quick internalization, minimum cell culture involvement, and low contamination risk. However, the delivery outcome of electroporation heavily depends on the treated cells such as their type, size, and orientation to the electric field, not ideal for highly heterogeneous blood samples. Herein, a new electroporation system was developed towards effective transfection to cells in blood regardless of their large diversity. By coupling replica molding and infiltration-coating processes, we successfully configured a three-dimensional electrode comprised of a polymer micropillar array on which carbon nanotubes (CNTs) are partially embedded. During electroporation, cells sag between micropillars and deform to form a conformal contact with their top and side surfaces. The implanted CNTs not only provide a robust conductive coating for polymer micropattern but also have their protruded ends face the cell membrane vertically everywhere with maximum transmembrane potential. Regardless of their largely varied sizes and random dispersion, both individual blood cell type and whole blood samples were effectively transfected with plasmid DNA (85% after 24 h and 95% after 72 h, or 2.5-3.0 folds enhancement). High-dose RNA probes were also introduced, which regulate better the expression levels of exogenous and endogenous genes in blood cells. Besides its promising performance on non-viral delivery routes to cell-related studies and therapy, the involved new fabrication method also provides a convenient and effective way to construct flexible electronics with stable micro/nano features on the surface.