Nanosecond time resolved measurement of membrane charging

Membrane charging is a necessary condition for subsequent membrane permeabilization. The time constant of membrane charging is on the order of 100 ns. Thus, measurements on the dynamics of membrane charging demand for fast diagnostic tools like pulsed laser fluorescence microscopy (PLFM) exhibiting a temporal resolution of 5 ns.

For measurement, the cell membrane is stained by a voltage sensitive dye. The cells, located between two micro electrodes on the microscope stage, are illuminated by a 5 ns long laser pulse during electric field pulse application. Membrane voltage changes due to membrane charging are indicated by intensity changes of the fluorescent light emitted from the dye molecules. Hyperpolarization results in higher emission intensity. At the depolarized cell hemisphere light intensity goes down. Current basic research deals with the dynamics of the transmembrane voltage during and after field exposure primarily targeting on increasing basic understanding of field induced permeabilization processes and on identifying energy-efficient treatment protocols for PEF processing.