Electroporation is a widely used non-viral technique for the delivery of molecules, including nucleic acids, into cells. Recently, electronic microsystems that miniaturize the electroporation machinery have been developed as a new tool for genetic manipulation of cells in vitro, by integrating metal microelectrodes in the culture substrate and enabling electroporation in-situ. We report that non-faradic SiO2 thin film-insulated microelectrodes can be used for reliable and spatially selective in-situ electroporation of mammalian cells. CHO-K1 and SH-SY5Y cell lines and primary neuronal cultures were electroporated by application of short and low amplitude voltage transients leading to cell electroporation by capacitive currents.
Fig: Cells on SiO2 thin-film capacitive microelectrodes. (a) Top view of the electroporation microchip including the cell culture plastic chamber glued on a ceramic package providing mechanical support and electrical contacts to the voltage generator. Scale bar: 1 cm. (b) Magnification of three octagonal microelectrodes out of sixty-four (organized in two linear arrays as shown in Fig) and with CHO-K1 cells growing in adhesion on top of them. Scale bar: 10 μm. (c, e) Drawing of the cell-chip interface and microelectrode structure (cross-section). The cell grows in adhesion on top of the SiO2 thin film from which it is separated by a narrow (i.e., in the range of a few tens of nanometers) cleft occupied by electrolyte. The highly conductive p-type silicon forms a capacitor together with the electrolyte and the oxide dielectric film separating the two. Insulation between microelectrodes is ensured by SiO2. (d) Equivalent electrical circuit of the cell-capacitive microelectrode (CME) system. The cell is described by two compartments of lipid membrane, one for the region of adhesion (with resistance RAM and capacitance CAM), the other one for the free portion exposed to the bulk electrolyte (RFM and CFM). Rcleft represents the resistance sealing the cleft from the bulk electrolyte. COx is the capacitance of the oxide and VS is the voltage applied to one face of the dielectric through the p-type Si. Accordingly, upon a change of Vs two potentials, ΔVF and ΔVA, develop across the two compartments of the cell membrane, eventually causing electroporation. VB: is the bias voltage applied to the bulk n-type silicon. (f) Stimulation settings. The bath electrolyte is kept at ground potential through a Ag/AgCl electrode and the bulk n-type Si at a bias potential VB. A voltage source, VS, modulates the p-type Si. A typical stimulation waveform consisting of a repetition of sawtooth waves is sketched at the bottom.
We demonstrate reliable delivery of DNA plasmids and exogenous gene expression, accompanied by high spatial selectivity and cell viability, even with differentiated neurons. Finally, we show that SiO2 thin film-insulated microelectrodes support a double and serial transfection of the targeted cells.
Maschietto, M., Dal Maschio, M., Girardi, S. et al. In situ electroporation of mammalian cells through SiO2 thin film capacitive microelectrodes. Sci Rep 11, 15126 (2021). https://doi.org/10.1038/s41598-021-94620-8