A programmable electroculture platform for direction-dependent alternating electric field delivery in cancer cell models.

Alternating electric fields (AEFs) at intermediate frequencies have been shown to disrupt mitosis and inhibit the proliferation of rapidly dividing tumor cells. Here, we developed a programmable in vitro AEF exposure system (0-500 kHz, 1-3 V/cm) featuring an electroculture platform compatible with standard 12-well tissue culture plates. The system enables investigation of cellular responses to fixed orthogonal field orientations and periodic polarization switching (PPS), defined as sequential alternation of the field direction between x- and y-axes at user-configurable intervals of 10-300 s Approach: Exposure conditions were characterized using a fully three-dimensional (3D) finite-element model to evaluate electric field distribution and specific absorption rate (SAR), while thermal simulations and experimental temperature measurements were used to assess thermal stability and non-thermal operation. Human glioblastoma (U-118 MG), neuroblastoma (SH-SY5Y), and non-malignant endothelial (HUVEC) cells were treated with AEFs (200 kHz, 3 V/cm) for 24-72 h. Cytotoxic effects were assessed using MTT assay, Live/Dead fluorescence staining, and Annexin V/PI imaging, followed by quantitative image analysis. Main results: AEF treatment induced significant, time-dependent reductions in cancer cell viability, reaching an approximately 80% reduction relative to controls in U-118 and SH-SY5Y cells after 72 h. Fluorescence imaging supported pronounced cytotoxic effects in malignant cells, whereas endothelial cells showed comparatively lower sensitivity. In the PPS mode evaluated after 24 h with a 300 s switching interval, viability in both cancer cell lines was further reduced by approximately 15% relative to the corresponding fixed-axis configurations. Significance: These findings show that, under the tested AEF exposure parameters, periodically switched field orientations can produce stronger cellular responses than fixed-orientation AEF delivery, including the first demonstration in SH-SY5Y cells. By integrating programmable directional switching into a standard 12-well electroculture format, the platform provides a configurable framework for future studies of field orientation and switching interval modulation.&#xD.