Lowering Series Resistance in Whole-Cell Patch Clamp Experiments using the Pushpen Patch Clamp Electrode

Abstract

Series resistance introduces voltage clamp errors in whole-cell patch clamp experiments and reduces the bandwidth of recordings by forming a low-pass filter with the cell membrane capacitance. The bulk of the series resistance is located at the pipette tip. During an experiment, cellular debris or membrane fragments can clog the tip and result in an increase in resistance. This effect is most notable when recording from small neural structures, as the pipette tip used is smaller than usual then. An increase in series resistance seriously limits the duration of recording and the reliability of the data recorded. The current procedure to lower series resistance is to apply gradual pulses of suction to clear the pipette tip of cellular debris or membrane, but this usually results in disruption of the gigaseal and ends the experiment. We introduce a new procedure for clearing clogged tips using our Pushpen patch clamp electrode.The Pushpen patch clamp electrode is a novel electrode for whole-cell patch clamp experiments, in which a linear motor moves a conical tungsten wire wound with Ag/AgCl wire, linearly inside the pipette. The tungsten wire tip can protrude from the pipette tip like a push pen, the procedure we call the “pushpen operation”. Using the pushpen operation, we obtain whole-cell configuration from cell-attached without applying suction but by impaling the cell membrane with the tungsten wire tip, while maintaining gigaseal. Currently we are working to demonstrate that we can lower series resistance during an experiment by using the pushpen operation to clear a clogged pipette tip and thus prolong the lifetime of experiments.