GE EPS 3501 XL User Manual page 24

5 Operation
24
During discontinuous electrophoresis, however, the voltage is not distributed evenly
across the gel. These gels have a region with low ionic strength that causes a high
electrical field strength. This region increases as the electrophoresis proceeds. This
means that the main part of the voltage is spread over a greater and greater distance
and a higher and higher power is tolerated. This is the reason why constant current is
chosen for such applications. It is, however, recommended to also limit the power as a
precaution against overheating the gel. The power supply will probably switch over to
limiting power at the end of the run due to increased voltage.
The crossing-over between different parameters controlling the electrophoresis
can be illustrated by IEF (isoelectric focusing) using carrier ampholytes. A graphical
representation of the changes in power, voltage and current that may occur during
a typical IEF run is given in Fig 9. Since the pK values of the carrier ampholytes and
the proteins are temperature dependent, IEF must be carried out at a constant
temperature. Therefore cooling of electrophoresis equipment and controlling by power
is recommended. The main part of the IEF is thus controlled by power (phase II). The
conductivity is gradually decreasing because the carrier ampholytes and sample will
loose their net charge during the build up of the pH gradient. Thus the current will
decrease and the voltage increase at constant power. During the early stage of the
formation of the pH gradient it is important to limit the current. Otherwise the gradient
will be irregularly shaped (phase I). The conductivity is not constant along the gel so it is
important to also limit the voltage. This means running at constant voltage for the last
phase (phase III) to prevent local overheating.
For detailed information about parameter values, always follow the gel supplier's
recommendations.
I
Fig 9. Changes in power, voltage and current during an IEF run.
Electrophoresis Power Supply EPS 3501 XL User Manual 18-1130-22 Edition AD
II
Power
Preset time
III IV
Alarm