Bio-Rad Whole Gel Eluter and Mini Whole Gel Eluter - Manual (Page 16)

4.3 Optimization Procedure
There are multiple factors that can affect the elution of a protein or mixture of proteins
from a slab gel (i.e. protein pI, buffer pH, molecular weight, gel porosity, gel thickness, etc.).
Therefore, for every new sample, change in elution buffer or running conditions, and when
making changes to the conditions of the initial preparative slab gel electrophoresis, we rec-
ommend doing the following optimization procedure.
1. Run a preparative gel with your sample. Make sure that the parameters used for this gel
are the same ones used for subsequent elutions (i.e. denatured or non-denatured gels and
gel thickness).
2. Follow Sections 3.1–3.2 for the setup of the Whole Gel Eluter, except excise the prepar-
ative gel so that the separated protein bands run perpendicular to the elution chambers
and not parallel to them.
3. Run the Whole Gel Eluter according to Section 3.3.
4. Eluted samples are harvested over a time course. On the first time point, turn off the
power supply and harvest one elution chamber with a disposable transfer pipet. Re-estab-
lish the current to the eluter after sampling.
5. Repeat step 4 for each time point until all of the chambers have been harvested. Then
assay the samples or total protein.
6. Plotting protein concentration versus sampling time will show the optimum elution time
for the buffer system and set running conditions.
Following every elution run, stain the gel and the membrane with Coomassie ® Blue. This
will indicate the completeness of the elution and whether proteins were lost due to membrane
binding.
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Section 5
Maintenance and Sterilization
5.1 Cleaning the Whole Gel Eluter
The Whole Gel Eluter should be cleaned after every use. The base plate, bottom elec-
trode, elution chamber core and upper plate/electrode should be washed with a laboratory
detergent (for example, Bio-Rad Cleaning Concentrate, catalog number 161-0722), then rinsed
thoroughly with distilled water. The lid with electrical leads should be wiped with a damp
cloth and allowed to dry thoroughly before use.
5.2 Sterilization of the Whole Gel Eluter
The Whole Gel Eluter is not autoclavable. The unit may be sterilized with 70% ethanol.
Do not attempt to sterilize the lid (ethanol will cause crazing of the plastics). The blotting
paper and cellophane (or dialysis) membrane may be autoclaved if desired.
5.3 Cleaning the Vacuum Harvester
The harvester and tube rack should be washed with a laboratory detergent, then rinsed thor-
oughly with water. The vacuum tubing and needle array should be thoroughly rinsed as well.
Turn the harvester lid upside down and fill it with water. This will allow water to flow through
the tubing and out the array. Dry the tubing by pulling air through the harvester while it is
attached to a vacuum source.
Note: Do not attempt to sterilize the Vacuum Harvester. Use autoclaved or pre-sterilized
transfer pipets if sterile harvesting is required.
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