Penetrating Nanoelectrodes for Stimulation of Single-Cells

Monday, 29 May 2017: 10:50
Grand Salon A - Section 4 (Hilton New Orleans Riverside)
K. Garde and S. Aravamudhan (North Carolina A&T State University)
In this paper, we report on the development of two nanoelectrode geometries, namely nanopillar and nanofin to interface with live neuronal cells. Electrical stimulus is delivered via penetrating nanoelectrodes in order to stimulate live PC12 cells. The paper also reports on the time dynamics of cell penetration process using the nanoelectrode. We show that (a) nanoelectrodes of optimized size and shape can penetrate into PC12 cells without any loss of cellular function and (b) electrical stimulus can be applied through the penetrating electrodes in order to stimulate or differentiate PC12 cells without the use of conventional growth factors, genetic manipulators, cytokines or other chemical reagents. The nanoelectrodes (nanopillars) are designed to overcome the trade-offs between electrode impedance and electrode size. Compared to conventional 3x3 array of 150 nm diameter nanowire electrodes, the current 50-150 nm nanoelectrodes have reduced electrode impedance by factor of at least 20 due to a large capacitance and small charge transfer resistance, while maintaining minimal cell damage. The fabrication of nanoelectodes starts from silicon on insulator (SOI) substrate, followed by a series of lithographic patterning, deposition and etching steps including focused ion beam (FIB) ion-milling. The precise placement of cells on the nanoelectrodes is a challenging process. After surface functionalization with silane coupling (uncoated Si) or thiol chemistry (Au nanoelectrodes), non-contact microprinting method using an inkjet printer was also used to place cells on the nanoelectrodes. For cell differentiation, pulsed DC stimuli with monophasic 100 ms square pulse of 100 mV/cm field strength and 1 Hz frequency are applied. In order to validate this differentiation process, gene expression markers and morphological changes are monitored. Figure 1 shows the dynamics of the cell penetration process over time, recorded at 5 minutes to 5 hours. Figure 2 shows fixed SEM images of differentiated cells being penetrated by the nanoelectrodes without any loss of cellular function. Additionally, it is evident (figure 3) that the nanoelectrodes are clearly assessing the interior of the cells’ cytosol. This image was taken after FIB milling of one half of the electrode-cell interface.