ECIS Electroporation and Wounding Module
The ECIS automated wound-healing assay offers many advantages over traditional methods:
- Completely automated
- Creates a precisely defined wound
- Highly reproducible
- Quantitative data
- Adsorbed proteins are not disrupted
- Entire assay accomplished without opening the incubator door
- Continuous, real-time migration measurement
ECIS can be modified such that electroporation and automated wound healing assays (cell migration assays) can be performed. In normal ECIS measurements, a current in the microampere range is used. This is undetected by the cells, and, in its measuring mode, ECIS essentially eavesdrops on cell behavior. When the current is boosted 1000 fold to the millliampere range, the resulting voltages across the cell membranes result in electroporation.
If this current is applied for only a few milliseconds, the cells recover, and it is possible to insert membrane-impermeable molecule (including DNA or RNA constructs) into the cytoplasm. Then the system can be returned to its non-invasive mode and the effects of the electroporation can be monitored in real time.
If the high current is applied for several seconds, cell death ensues due to severe electroporation and possible local heating effects. Only the cells on the small active electrodes are killed, and then the instruments can be used to monitor cell migration to repopulate the wounded area.
The ECIS wound is very well defined, as it includes only those cells on the 250 micrometer diameter electrode. Death can be verified both with the ECIS measurements and with vital staining. Typical ECIS data involving this assay is shown in the figure below. Here BSC1 cells were first grown as complete monolayers, and the impedance traces from four confluent wells can be seen on the graph. At the arrow, an elevated field was applied to two of the wells, wounding the cells on the small electrode and causing the impedance to drop to that of an open electrode. Over time these two traces return to control values, as the healthy cells outside of the small electrode migrate inward to repopulate the wounded area and replace their dead cohorts (healing). These types of data are highly reproducible and respond to culture conditions.
Below are scanning electron micrographs of the ECIS electrode with NRK cells during a wound healing experiment:
The "Electric Fence" Cell Migration Assay
Applied Biophysics has developed a novel impedance-based technique called “The Electric Fence” to measure the rates of cell migration.
The Electric Fence differs from the Wounding Assay in that it prevents the cells from actually growing on the electrode while a confluent layer develops around the electrode. When the "electric fence" is activated, a series of high field electric pulses are applied which prevent the cells from attaching and spreading onto the measurement electrode. When the electric fence is turned off, the cells in the surrounding confluent layer migrate into the open space left by the electric fence.
The progress of the migration into this open space is very precisely monitored and from this data a migration rate is calculated. The key advantage of the Electric Fence over the Wounding Assay is the surface which the cells migrate onto has not been modified in any manner by previous cell growth, should a protein layer have been added prior to the experiment.