Bio-Rad Model 111 Mini IEF Cell - Manual (Page 14)

3.7 Sample Application
Sample application is most conveniently accomplished using the included Sample Templates.
1. Place the template on top of the polymerized gel. The colored portion of the template should
coincide with the longer dimension of the gel. The application position for the sample varies
(see Section 3.8) but allow 1 cm at both the top and bottom of the gel for the gel to contact the
electrodes.
2. Apply samples using a pipettor capable of delivering between 0.5 µl and 2 µl. Volumes above
2 µl are not generally recommended for use with the sample template.
3. Allow samples to diffuse into the gel for 5 minutes.
4. Carefully remove the template from the gel.
Note: For larger samples, one may custom-form application strips from filter paper or use
Bio-Rad’s Sample Application Pieces (catalog number 170-4257). Apply the sample to the
filter paper and then place the filter paper at the point of application on the gel. Allow the
sample to diffuse into the gel for 5 minutes. The disadvantage of this method is that some
proteins may not be completely eluted from the strip to the gel.
3.8 Position of Application
There are no fixed rules regarding the positioning of the sample on the gel. Samples should
not, in general, be applied to areas where protein bands are expected to focus. To protect the
proteins from extreme pH exposure, the samples should not be applied closer than 1 cm from
either electrode. The best strategy for a new protein is to make three points of application, one at
each end and one near the middle of the gel, and observe the resulting focusing pattern.
Section 4
Running the Gel
4.1 Set Up Procedure
1. Slide the lid of the Model 111 Mini IEF Cell toward the electrode plugs to remove it.
2. Remove the Graphite Electrodes from the cell and rinse them with distilled water to remove
traces of acrylamide from the previous run. The electrodes may be gently wiped with a
laboratory tissue if necessary. Place the electrodes back into the cell.
3. Lightly moisten the graphite electrodes with water. Turn the gel with the adsorbed samples
upside-down and place it directly on top of the electrodes. Position the gel carefully the first
time, as re-positioning may damage the gel surface. Do not remove the glass plate from the
gel/gel support backing because its weight insures good contact between the gel and the
electrodes.
4. Carefully slide the lid back onto the Model 111 Mini IEF Cell. Plug the power cables of the
cell into a power supply able to generate 500 V constant voltage. The cell is now ready to
separate samples.
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