Bio-Rad PROTEAN II XL Cell User Manual
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6. Prepare the stacking gel monomer solution. Combine all reagents except APS and TEMED, and
deaerate under vacuum at least 15 minutes.
7. Dry the area above the separating gel completely with filter paper before pouring the stacking gel.
8. Place a comb in the gel sandwich, and tilt it so that the teeth are at a slight (~10°) angle. This will prevent
air from being trapped under the comb teeth while pouring the monomer solutions.
9. Add APS and TEMED to the degassed monomer solution, and pour the solution down the spacer
nearest the upturned side of the comb. Pour until all the teeth have been covered by solution. Then
properly align the comb in the sandwich, and add monomer to fill completely.
Generally, an overlay solution is not necessary for polymerization when a comb is in place.
10. Allow the gel to polymerize 30-45 minutes. Remove the comb by pulling it straight up slowly and gently
11. Rinse the wells completely with distilled water. The gels are now ready to be attached to the central
cooling core, the sample loaded, and the gels run.