Dilutions of stock were prepared in cell culture medium with required concentrations.
Cells (5 × 10 per well) seeded in 96-well plates were treated with required concentrations of the drug.
After a stable baseline electrical signal was obtained, the DMMP vapor with the required concentration was introduced, and all sensing measurements were carried out at room temperature.
Briefly, cells were stimulated with the required concentration of NECA for 5 min in fresh serum-free medium.
For the ligand-binding experiments, the cells were incubated with the required concentration of fluorescent ligand for 10 min at 37°C before imaging.
ECV304 cells (10 cells per well) were added to the matrigel layer together with the required concentration of XAA analogues in a final volume of 1 ml supplemented culture medium.
Sunitinib (20 mmol/L in DMSO, Pfizer, Cappele a/d IJssel, The Netherlands) was diluted in 0.9% saline, as a control 0.9% saline with the required concentration of DMSO was used.
For the diploid cdc28-as1 (F88G) strain, cells were grown to exponential phase in YPD, harvested, and the medium replaced with YPD plus the required concentration of inhibitor, and returned to 30C with samples taken at 30 minutes, 1 hour, 2 hours and 3 hours thereafter.
The reduced selenols were diluted with degassed phosphate buffer to the required concentration (9.9 86.4 μM), with the concentrations determined using their corresponding molar absorption coefficients.
