Patch Clamp - Basic Technique

Basic Technique

Patch clamp recording uses, as an electrode, a glass micropipette that has an open tip diameter of about one micrometre, a size enclosing a membrane surface area or "patch" that often contains just one or a few ion channel molecules. This type of electrode is distinct from the "sharp microelectrode" used to impale cells in traditional intracellular recordings, in that it is sealed onto the surface of the cell membrane, rather than inserted through it. In some experiments, the micropipette tip is heated in a microforge to produce a smooth surface that assists in forming a high resistance seal with the cell membrane. The interior of the pipette is filled with a solution matching the ionic composition of the bath solution, as in the case of cell-attached recording, or the cytoplasm for whole-cell recording. A chlorided silver wire is placed in contact with this solution and conducts electric current to the amplifier. The investigator can change the composition of this solution or add drugs to study the ion channels under different conditions. The micropipette is pressed against a cell membrane and suction is applied to assist in the formation of a high resistance seal between the glass and the cell membrane (a "gigaohm seal" or "gigaseal," since the electrical resistance of that seal is in excess of a gigaohm). The high resistance of this seal makes it possible to electronically isolate the currents measured across the membrane patch with little competing noise, as well as providing some mechanical stability to the recording.

Unlike traditional two-electrode voltage clamp recordings, patch clamp recording uses a single electrode to record currents. Many patch clamp amplifiers do not use true voltage clamp circuitry but instead are differential amplifiers that use the bath electrode to set the zero current level. This allows a researcher to keep the voltage constant while observing changes in current. Alternatively, the cell can be current clamped in whole-cell mode, keeping current constant while observing changes in membrane voltage.