The patch clamp technique is a laboratory technique in electrophysiology that allows the study of single or multiple ion channels in cells. The technique can be applied to a wide variety of cells, but is especially useful in the study of excitable cells such as neurons, cardiomyocytes, muscle fibers, and pancreatic beta cells. It can also be applied to the study of bacterial ion channels in specially prepared giant spheroplasts.
The patch clamp technique is a refinement of the voltage clamp. Erwin Neher and Bert Sakmann developed the patch clamp in the late 1970s,the voltage-clamp technique and for the first time resolved single channel currents across a membrane patch of a frog skeletal muscle. They were also honored with the Nobel Prize in Physiology and Medicine (in 1991). The next breakthrough was the invention of the giga seal by Ernst Sakmann in 1980 which immensely improved the signal-to-noise ratio and allowed the recording of even smaller currents.Electrophysiology, pioneered in special biophysical laboratories, now expanded to basic biological and medical research and became one of the most important tools for the investigation of the behavior of single cells or whole cellular networks in the nervous system.
Principle
The patch-clamp technique allows the investigation of a small set or even single ion channels. It is thus of special interest in the research of excitable cells such as neurons, cardiomyocytes and muscle fibers.
A single ion channel conducts around 10 million ions per second. Yet the current is only a few picoamperes. Recording currents in this order of magnitude is quite challenging not only for the researcher, but also for the equipment. In principle, thin glass or quartz pipettes with a blunt end are sealed onto the membrane.
Suction is applied to aid the development of a high-resistance seal in the gigaohm range. This tight seal isolates the membrane patch electrically, which means that all ions fluxing the membrane patch flow into the pipette and are recorded by a chlorided silver electrode connected to a highly sensitive electronic amplifier. A bath electrode is used to set the zero level.
To prevent alterations in the membrane potential, a compensating current that resembles the current that is flowing through the membrane is generated by the amplifier as a negative feedback mechanism.
The membrane potential of the cell is measured and compared to the command potential. If there are differences between the command potential and the measurement, a current will be injected. This compensation current will be recorded and allows conclusions about the membrane conductance. The membrane potential can be manipulated independently of ionic currents and this allows investigation of the current-voltage relationships of membrane channels.
