Hoefer SE400 - Manual (Page 20)

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3
If combs are in place, skip to step 4.
If no combs are in place, overlay the resolving gel
with a thin layer of water-saturated n-butanol, water,
or diluted gel buffer to prevent exposure of the top
surface of the gel solution to atmospheric oxygen.
Slowly deliver the overlay solution from a glass
syringe fitted with a 22-gauge needle. Apply the
solution near the spacer and allow it to flow across
the surface unaided.
4
Allow the gel to polymerize for a minimum of
one hour.
2.2.2 Stacking gel
Pour the stacking gel before removing the
sandwich from the gel caster. Stacking gel
resolution is optimal when prepared just before
electrophoresis.
1
Remove the overlay by rinsing the top of the gel
several times with distilled water. Invert the caster
to drain. To ensure a seamless contact between the
resolving and stacking gels, remove residual liquid by
blotting one corner with a lint-free tissue.
2
Calculate the stacking gel monomer solution volume.
3
Prepare the stacking gel monomer solution, deaerate
it, and add catalyst and initiator. Pour the stacking
gel onto the resolving gel with a disposable or Pasteur
pipet to a level about 2 mm from the top of the
glass plate.
4
Introduce a comb (at a slight angle) into the sand-
wich, taking care not to trap air under the teeth. Allow
a minimum of one hour for the gel to polymerize.