Bio-Rad Model 111 Mini IEF Cell User Manual
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4.2 Run Conditions
1. Focusing is carried out under constant voltage conditions in a stepped fashion. Begin focusing
at 100 V for 15 minutes.
2. Increase voltage to 200 V for 15 minutes.
3. Finally, increase the voltage to 450 V for an additional 60 minutes.
Note: Step increases of voltage are necessary to prevent overheating and subsequent
dehydration of the gel. Failure to follow this procedure will result in poor resolution. One
advantage of using Bio-Rad’s PowerPac 3000 or Model 1000/500 power supply is that either
power supply can be programmed to carry out this stepped increase in voltage automatically.
4. Typical running conditions for various ranges of Bio-Lyte ampholytes in the Model 111 Mini
IEF Cell are shown in Table 4.1. Any gross variation from these values could indicate
problems associated with polymerization, buffer preparation, etc.
Table 4.1 Typical Running Conditions
Bio-Lyte pH
Current/Power
Current/Power
Current/Power
Range
at 100 V
at 200 V
at 450 V
3-10
5-6 mA/0.5-0.6 W
5-6 mA/1 W
4 mA/2 W
4-6
3 mA/0.3 W
3-4 mA/0.6-0.8 W
4-5 mA/2 W
5-7
3 mA/0.3 W
3-4 mA/0.6-0.8 W
4-5 mA/2 W
6-8
3 mA/0.3 W
3-4 mA/0.6-0.8 W
4-5 mA/2 W
7-9
3 mA/0.3 W
3-4 mA/0.6-0.8 W
4-5 mA/2 W
8-10
10 mA/1 W
20 mA/4 W
4 mA/2 W
5. A good way to monitor the progress of an isoelectric focusing experiment is to observe the
migration of visible marker proteins of known pI to their isoelectric points. The IEF Standards
(catalog number 161-0310) provide eight natural proteins, including four visible proteins that
are clearly discernable during an IEF run. These markers are useful with all non-denaturing
IEF buffer systems.
6. As the focusing nears completion, the current will decrease from the values listed in the 450 V
column. This is a general indication that the focusing is near completion.
Note: The Model 1000/500 Power Supply measures current to the microampere range. This
expanded monitoring of the changes in current allows better estimation of the time the current
stabilizes and the experiment is finished.
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