Poly l lysine 0.01

The HEK293H cell line was obtained from ThermoFisher Scientific Invitrogen (Waltham, MA) and cultured with a DMEM/high glucose medium (GE Healthcare Life sciences, Piscataway) supplemented with fetal bovine serum 10% (ThermoFisher Scientific Gibco, Waltham, MA) in humidified incubator with 5% CO₂ at 37 °C. Cells were seeded at a density of 1 × 10⁵ per well. To record the transmembrane potential, cells were placed on a cover glass treated with poly-L-lysine 0.01% (Sigma-Aldrich). For the SICM imaging, cells were seeded on a polydimethylsiloxane (PDMS) substrate in a petri dish with 2 mm of thickness. The PDMS substrate was coated with collagen hydrogel matrix (5 μg/cm²) to improve the cell adhesion.^{29 (link)} If not mentioned otherwise, the HEK293H cells were treated with 10 μM K₂CrO₄ in extracellular solution for 90 min. For the fixed cell experiments by SICM, two batches of chromate treated cells were fixed with 4% paraformaldehyde for 10 min. Before patch-clamp or SICM experiments, the treated cells were rinsed two times with the extracellular solution.

A Bruker Multimode 8 was used in tapping-mode with an J-type scanner, Nanoscope VI controller, Nanoscope v614r1 control software (Veeco, Cambridge, U.K.) and a silicon tapping tip (NSG01, NTI-Europe, Apeldoorn, The Netherlands) of 10 nm curvature radius, mounted on a tapping mode silicon cantilever with a typical resonance frequency 283–374 kHz and a force constant of 12–103 N/m (Bruker OTESPA, U.K.). Images were taken in air, by depositing 40 μL of the sample on a freshly cleaved mica surface (Agar Scientific, Essex, U.K.) coated with poly-L-lysine 0.01% (Sigma-Aldrich, U.K.) and allowed to adsorb for 2 min. Excess unbound material was removed by washing with Milli-Q H₂O and then drying in air; this step was repeated once. Size distributions were carried out using ImageJ software to measure the lateral dimension of individual graphene sheets, by counting more than 100 sheets. Sheet thickness was determined from the AFM height profiles.