Hoefer SE300 miniVE - Manual (Page 17)

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Warning! Acrylamide is a
neurotoxin. Always wear gloves
and observe all laboratory
safety procedures.
Note: Approximately 10 ml of
monomer solution is required
to cast one 1-mm thick gel.
A 1-cm stacking gel below the wells
Fill to 3 cm below the top of the rectangular
glass plate. Overlay each gel with a thin layer of
water-saturated n-butanol, water, or diluted gel
buffer to prevent exposing the monomer solu-
tion to oxygen. Use a glass syringe fitted with a
22-gauge needle to apply 100 µl of the overlay
solution slowly to one side of the sandwich, near
the spacer. Allow the solution to flow across the
surface unaided.
After polymerization

Allow a minimum of one hour for the gel to polymerize.

If a comb is in place, remove it by carefully pulling
on the comb while gently rocking it back and forth to
break the vacuum. Rinse the wells with electrophore-
sis buffer to remove any unpolymerized acrylamide.
If an overlay was applied, rinse the sandwich several
times with double-distilled water to remove it. Invert
the module to drain.
To ensure seamless contact between the resolving and
stacking gels, remove residual liquid by blotting one
corner of the gel with a lint-free tissue.
Casting the stacking gel

Prepare the stacking gel monomer solution.

Deaerate the stacking gel monomer solution, add
catalyst and initiator and then pour.
Use a pipette to deliver the solution into one corner of
the plate, taking care not to trap any bubbles.