Laser-Activated Plasmonic Substrates for Intracellular Delivery

Abstract

This thesis explores the use of laser-activated plasmonic pyramids for intracellular delivery. For our first study, we designed pyramids on a substrate with a nano-aperture on top to investigate computationally the near-field enhancement from femtosecond laser excitation. Simulations showed a 100-fold enhancement of intensity at the aperture of the pyramid. We fabricated substrates using template-stripping, and use Nearfield Scanning Optical Microscopy to show local field enhancement on nano-aperture. For our second study, we designed and fabricated large-area (14 x 14 mm), photolithography-based, template-stripped plasmonic substrates that are nanosecond laser-activated to form transient pores in cells for cargo entry. In this iteration of the design, the aperture on the pyramids was removed to make the fabrication more consistent from batch to batch. Temperature simulations show the pyramid apex reaching bubble formation temperatures. We used flow cytometry to characterize the delivery efficiency of cargos. This technique offers additional desirable features: spatial selectivity, reproducibility, minimal residual fragments, and cost-effective fabrication. The rest of the thesis builds a biophysical understanding of cellular structural changes and pore formation in cells using fluorescence microscopy, which is important for the development of safe therapeutic applications. In the final section of the thesis we show proof-of-principle results with experiments using blood stem cells, and also using a more affordable continuous laser. The research presented in this thesis supports the development of safer genetic and viral disease therapies as well as novel research tools for fundamental biological research that rely on effectively delivering molecules to millions of living cells.