Bosch etching for the creation of a 3D nanoelectroporation system for high throughput gene delivery

In order to create a high-throughput electroporation based cell transfection system, it is required that each cell has localized delivery and minimal membrane damage to ensure optimal transfection and longevity post-biomolecule delivery. To meet these requirements, a three-dimensional (3D) nanochannel device was fabricated on a Si platform due its ease of etching, wide industrial availability, and mechanical stability. The device is designed to shoot desired biomolecules into a seated array of target cells to achieve the high-throughput of bulk electroporation, but with greatly reduced cell mortality. To accomplish this, a wafer-scale Bosch etching process was optimized to etch a 3D array of channels consisting of larger microchannels feeding into smaller nanochannels that cells are ultimately seated on for transfection. The microchannel array consists of 50 μm wells spaced 50 μm apart, which are etched from the "back side." The wafer is then flipped over to etch the smaller 650 nm channels on the "front side." In the creation of the 3D silicon device, other feature sizes were explored, and their Bosch etching was characterized for comparison. The results show that when etching samples with the same feature sizes, but different densities, there was no relation between feature density and etch rate for our recipe. However, when etching features, or more specifically, circular channels of different sizes (650 nm-150 μm), the results show a positive correlation with etch rate (1.10-4.06 μm/min). Standard deviations indicate very uniform etching with an average value of 0.1 μm/min across all etches. After optimization, the 3D Si device was tested to ensure successful cell seating and transfection via electroporation, using fluorescence as the tool of evaluation. Fluorescent imaging (postelectroporation) indicates a transfection efficiency of approximately 70% with a cell viability of roughly 90%.