To perform intracellular delivery with NES, a total of 50,000 to 100,000 cells of interest were cultured on the NS platform within a 5-mm plastic tube for 0.5 to 7 days with the respective culture media. The recipes for each cell culture medium are listed in table S1. The cell culture devices were placed in a 48-well plate for incubation (5% CO2, 37°C). After incubation, the cell culture devices were placed on an ITO electrode with 2- to 5-μl droplets of the delivery reagent at the specified concentration. A platinum electrode (0.5 mm in diameter) was immersed in the cell culture. Square pulses of 20 Hz and pulse duration of 20 V and 200 μs were applied between the platinum electrode and ITO electrode for 20 s. The positions of the cathode and the anode were determined by the charge of the delivery molecules. For example, if the delivery molecules were charge negative, then the platinum electrode served as the anode. After delivery, the cell culture tube was returned to the 48-well plate for further incubation. When delivering fluorescently labeled molecules, the cells were washed three times with 1× PBS to remove excess molecules preceding analysis.

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