Gene electrotransfer is one of the main non-viral methods for intracellular delivery of plasmid
DNA, wherein exposure of biological cells to pulsed electric field induces electroporation, that
is a transient increase in cell membrane permebility to exogenous molecules. By localizing the
electric field on small parts of the cell membrane using nanostructured geometries such as
nanostraws, nanochannels, and nanopores, the efficiency of gene electrotransfer and cell
viability can be significantly increased. Disadvantages of such nanofabricated devices are that
they are not widely available and require nanofabrication expertise and access to cleanroom. It
has recently been shown that highly efficient electrotransfer of plasmids can also be achieved
using commercial polycarbonate membranes with nanopores of 0.05 μm in radius. Therefore,
in this thesis I study and design an electrotransfer approach based on commercially available
cell culture inserts with porous membrane from polyethylene terephthalate (PET). First, I use
numerical modeling at the single cell level to determine electric pulse parameters, size of pores
in the porous membrane, and other factors expected to result in successful gene electrotransfer.
Based on the numerical results, I design a simple device that contains an insert with porous
membrane containing pores with radius of 0,2 µm, 0,5 µm or 1,5 µm, a multiwell plate, and a
pair of wire electrodes. I numerically model the device to improve the placement and
configuration of the electrodes, in order to achieve a homogenous voltage along the entire
porous membrane. I then test the device using three mammalian cell lines and assess the
transfection efficiency and cell viability. The results are comparable to those obtained in
previous studies using classical gene electrotransfer, where cells in suspension or adhered to
surface are placed between a pair of parallel electrodes and exposed to homogenous electric
field. However, the porous membranes enable application of low-voltage pulses, which do not
require the use of expensive high-voltage electroporators. Finally, I discuss further research
directions to assess the influence of porous membrane properties, such as thickness, porosity,
and pore radius on transfection efficiency and cell viability.
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