I only have experience with Axoclamp 2B from when I was a student but now I have to use Axopatch 200B for perforated patch and there isn't much in the manual about perforated patch. My question is about "setting the cell up" before a perforated patch experiment, e.g. capacitance compensation, optimising the voltage clamp. When I used the Axoclamp 2B I used discontinuous single electrode voltage clamp (dSEVC) mode and an oscilloscope to monitor the switch clamp. How does this process differ with the Axopatch 200B?
Dear Fluer,
In my practice, there is no special needs for the “setting the cell up” using 200B in perforated mode (I assume you are going to use whole-cell voltage clamp). All procedures for regular whole-cell recordings are applicable in perforated mode.
Since you will need monitoring the perforation process (establishing a cell capacitance), at the beginning it is prerequisite to switch off all compensations. When you reach a steady state (with antibiotics usually it will take 3 – 5 min, depending on cell type), then it would be necessary to take any means to compensate access (series) resistance (this feature looks advanced in Axopatch 200B), especially if you will measure significant whole-cell membrane currents. Such value of resistance, as a rule, is significantly higher compared to that obtained in conventional whole-cell. However, when access resistance is appropriate (at measured current, it would not provide meaningful drop in command potential) and capacitive transitions do not critically interfere with measured currents, I prefer avoid compensation of cell capacitance, since non-compensated capacitive currents can facilitate monitoring the quality of perforation, which may not be consistent during experiments. Alternatively, 200B has a switch to turn on/off “whole cell parameters”, which can be operatively used to exit/enter whole-cell compensations.
Dear Alexey,
Thank you very much for your answer. I will see how we get on and may come back to you if there are any issues, if that is ok!