Visualization and characterization of membrane lesions

In this study, real-time imaging was used to visualize individual electropores and demonstrate their persistence in electroporated cells. HEK 923 cells were cultured on Indium Tin Oxide (ITO)-coated coverslips and loaded with the Cal520 fluorescent indicator for visualization. Real-time imaging was performed in Total Internal Reflection Fluorescent (TIRF) mode. Electrophysiological manipulations were applied using a patch clamp in voltage clamp mode, set in a whole-cell configuration. The protocol included a command voltage of 0 mV interrupted by a -50 mV pulse for 50 ms and a -400 mV pulse for 1 ms at frame 91, returning to -50 mV for 50 ms. Additionally, 0.4 seconds into the second and third stacks, the command voltage was changed from 0 mV to -50 mV for 50 ms. The study was supported in part by NIH 15R21EY034803 and NIH 1R21EY034258.