Development of a carboxyl-terminated Indium Tin Oxide electrode for improving cell adhesion and facilitating low noise, real-time impedance measurements

The working electrode's surface property is crucial to cell adhesion and signal collection in electric cell-substrate impedance spectroscopy (ECIS). To date, the indium tin oxide (ITO) based working electrode is of interest in ECIS study due to its high transparency and biocompatibility. Of great concern is the impedance signal loss, distortion, and data interpretation conflict profoundly created by the movement of multiple cells during ECIS study. Here, a carboxyl- terminated-ITO substrate was prepared by stepwise surface amino silanization, with N-Hydroxy succinimide (NHS), and (1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide hydrochloride) (EDC) treatment, respectively. We investigated the stepwise changes in the property of the treated ITO, cell-substrate adhesion, collective cell mobility, and time course of change in absolute impedance from multiple Chinese hamster ovary (CHO) cells ((Δt-Δ|Z|)_CELLS). The carboxyl-terminated ITO substrate with a surface roughness of 6.37 nm shows enhanced conductivity, 75% visible light transparency, improved cell adherence, reduced collective cell migration speed by ~2 fold, and diminished signal distortion in the ((Δt-Δ|Z|)_CELLS). Thus, our study provides an ITO surface-treatment strategy to reduce multiple cell movement effects and to obtain essential cell information46 from the ECIS study of multiple cells through undistorted (Δt-Δ|Z|)_CELLS.