Cultivation of cells under strong ac-electric field—differentiation between heating and trans-membrane potential effects

Adherently growing mouse fibroblasts were cultivated for days in highly conductive culture media between micro-fabricated electrodes under strong ac-electric fields. Miniature electrodes have improved heat dissipation which allows the use of media with conductivity of more than 1 S/m at field strengths of up to 100 kV/m. Cell division rates, cell motility, cell viability and physiological parameters such as vesiculation were monitored and the actin and β-tubuline structures of cyto-skeleton were imaged by laser scanning fluorescence. The specific effects of polarisation could be differentiated from unspecific effects such as heating. We estimated the real field strength acting on cells. In the kHz-range, field application was clearly limited by membrane dielectric breakdown while temperature increases were less than 3°C. In the MHz-range, much stronger fields could be applied and heating became the limiting factor. Above an induced trans-membrane potential of 130–150 mV cells no longer proliferated under prolonged field application. In the MHz-range (above 5 MHz) cells could be exposed to surprisingly high field strengths (40 kV/m) for days. Therefore, there is a frequency window (up to several 100 MHz) which can be used for cell positioning, manipulation and characterisation techniques without significant loading of cells.