The patch clamp technique allows for high-resolution recording of ion channel currents. It involves using a glass pipette to form a high-resistance seal with a cell membrane, isolating a small portion of the membrane. This enables measurement of the electric current passing through individual or small groups of ion channels as voltages are varied. The technique was developed in the late 1970s and has provided insights into the functions of many ion channels in excitable cells like neurons.

2. Presented by
Dilruba Afrin
Course : GEB-410
Dept. of Genetic Engineering &
Biotechnology
University of Rajshahi

3. Introduction
What is patch clamping?
Why used patch clamp?
Basic principle
Types and configurations
Application
Limitations

4. The patch clamp technique is a refinement of the voltage clamp.
Erwin Neher and Bert Sakmann developed the patch clamp in the
late 1970s and early 1980s . Neher and Sakmann received the Nobel
Prize in Physiology or Medicine in 1991 for this work.
 The method used by Neher, Sakmann & Steinbach in1978 and
subsequent refinements by Hamill ,Marty, Neher,Sakmann &
Sigworth in 1981 have led to techniques for high resolution recording
of current in excised membrane patches in addition to those that
remain cell-attached.

5.  Patch clamping is a widely applied electrophysiological technique
for the study of ion channels; membrane proteins that regulate the
flow of ions across cellular membranes and therefore influence the
physiology of all 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.

6.  Provides access to the inside of the cell.
• Can insert an electrode into the cell
• Can change the intracellular fluid
 Creates a seal impermeable to ion flow.
• High electrical resistance
 Allows to measure current through ion channels vs.
voltage,time,temperature,menthol,concentration etc.

7.  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.
1. In principle, thin glass or quartz pipettes with a blunt end are
sealed onto the membrane (Figure 2).
2. Suction is applied to aid the development of a high-resistance
seal in the gigaohm range.

8. 3. 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.
4. 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 (Figure 1).
5. 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.

9. 6. This compensation current will be recorded and allows
conclusions about the membrane conductance.
7. The membrane potential can be manipulated
independently of ionic currents and this allows
investigation of the current-voltage relationships of
membrane channels.

10. Fig.1 : General principle of patch-clamp
recordings. A glass pipette
containing electrolyte solution is
tightly sealed onto the cell
membrane and thus isolates a
membrane patch electrically.
Currents fluxing through the
channels in this patch hence flow
into the pipette and can be
recorded by an electrode that is
connected to a highly sensitive
differential amplifier. In the
voltage-clamp configuration, a
current is injected into the cell
via a negative feedback loop to
compensate changes in
membrane potential. Recording
this current allows conclusions
about the membrane
conductance.

11. Fig. 2: A phase contrast image of a patch pipette attached to
the membrane of a cultured murine hippocampal neuron.

12. Several variations of the basic technique
can be applied, depending on what the
researcher wants to study –
 Cell-attached or on-cell patch
 Inside-out patch
 Whole-cell recording or whole-cell
patch
 Outside-out patch
 Perforated patch
 Loose patch
 Automatic patch clamping
Diagram showing
configurations of the
patch clamp technique

13.  For the evaluation of antiarrhythmics agents.
 Used for isolated ventricular myocytes from Guinea pigs to study a
cardio selective inhibition of the ATP sensitive potassium channel.
 To identify multiple types of calcium channels.
 To measure the effect of potassium channel openers.
 Used in molecular biology.
 Voltage clamp studies on sodium channels.
 Used to investigate a wide range of electrophysiological cell properties.
 Measurement of cell membrane conductance.

14.  Imparting skillful training performance and recording in during
single channel recording.
 Cost of process is expensive.
 Time consuming.
 Number of samples required is more at times.
 Chance of membrane distortion.

15. Thanks to all