Bio-Rad PROTEAN® IEF Cell Accessories - Manual (Page 15)

Section 4
Running Preset Methods
The PROTEAN IEF Cell contains three preset methods to be used as a starting point for
new samples and novice users to help determine optimal focusing conditions. Each of the
three methods is based on the manner in which the voltage is ramped in step 2x (the slope to
reach the maximum voltage). In all preset methods the maximum applied voltage is based
on the length of the IPG strip (4,000 V/7 cm; 8,000 V/11 cm; 10,000 V/17 cm). The program
will prompt the user for the IPG strip size. Each of these methods provides an optimum
maximum field strength of ~600 V/cm of IPG strip. The pre-set methods have a 50 µA
limit/IPG strip. If a different current limit is needed a customized method can be programmed
using the NEW METHOD option.
Fig. 5. Preset method voltage over time.
Step Parameters
R - Rehydration (*optional) Passive, or Active at 50 V
S1 - Conditioning Step 250 V for 15 minutes. A low voltage is applied
in Step 1 to remove salt ions and charged
contaminants.
S2 - Voltage Ramping Slow [ ], Linear [ ] or Rapid [ ]. When the
conditioning step is complete, the voltage
ramping step increases the voltage to the
maximum voltage in one of three ways. In all
three cases the current will not exceed the
50 µA/strip limit. See Section 2.2 for details.
S3 - Final Focusing A time is programmed to complete the focusing
process once the maximum voltage is reached.
If the maximum voltage is not reached at the
completion of step 2, the voltage will increase
to the maximum voltage using the rapid ramping
method. The current limit will not exceed
50 µA/strip.
Voltage
IPG Strip
4,000 V 7 cm
8,000 V 11 cm
10,000 V 17 cm
S4 - Hold step (optional) 500 V is maintained until the run is stopped
to prevent diffusion of focused proteins or
over-focusing artifacts.
*Rehydration step can be included in the preset method program or performed separately.
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SLOW RAMP [ ]
LINEAR RAMP [ ]
RAPID RAMP [ ]
S4
S3
S2
S1
250 V
500 V
50 V
Max V
VOLTAGE
R
TIME/VH