Bio-Rad Mini-PROTEAN® Tetra Cell - Manual (Page 12)

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iv. Immediately overlay the monomer solution with water or
t-amyl alcohol.
Note: If water is used, add it slowly and evenly to prevent mixing.
Do not overlay with butanol or isobutanol.
v. Allow the gel to polymerize for 45 min to 1 hr. Rinse
the gel surface completely with distilled water. Do not leave
the alcohol overlay on the gel for more than 1 hr because
it will dehydrate the top of the gel.
Note: At this point the resolving gel can be stored at room
temperature overnight. Add 5 ml of 1:4 dilution of 1.5 M
Tris-HCl, pH 8.8 buffer (for Laemmli system) to the resolving gel
to keep it hydrated. If using another buffer system, add 5
ml 1x resolving gel buffer for storage.
vi. Prepare the stacking gel monomer solution. Combine all
reagents except APS and TEMED. Degas under vacuum for
at least 15 min.
vii. Dry the top of the resolving gel with filter paper before
pouring the stacking gel.
viii. Add APS and TEMED to the degassed stacking gel
monomer solution and pour the solution between the glass
plates. Continue to pour until the top of the short plate is
reached.
b. Continuous Polyacrylamide Gels
i. Prepare the monomer solution by combining all reagents
except the APS and the TEMED. Degas under vacuum for
15 min (refer to section 4 for gel formulations).
ii. Add APS and TEMED to the degassed monomer solution
and pour the solution between the glass plates. Continue to
pour until the top of the short plate is reached.
iii. Insert the desired comb between the spacers starting at the
top of the spacer plate, making sure that the tabs at the
ends of each comb are guided between the spacers. Seat
the comb in the gel cassette by aligning the comb ridge with
the top of the short plate.
iv. Rinse the casting frame(s) and stand with distilled,
deionized water after use.