Hoefer SE215 - Manual (Page 9)

2
Deaerate the monomer solution and add the initiator
and catalyst just prior to pouring the gel. Fill the solu-
tion into only one sandwich, allowing the groove in the
bottom plug to distribute the solution evenly between
all sandwiches. (Pouring into more than one sandwich
could introduce air pockets.) Watch the solution level
through the faceplate. See below for the appropriate
solution level according to the application.
No stacking gel
Fill with solution to just below the top of the notched
plate. If air pockets form, remove them with a pipette or
syringe. Introduce a comb (at a slight angle) into each
sandwich, taking care not to trap air under the teeth.
Stacking gel
Fill with solution to 3 cm below the top of the rectan-
gular glass plate. This height allows 1 cm of stacking
gel below the wells. Pour the gel and apply an overlay.
(See step 3.) After the gel is set, prepare the stacking
gel as described below.
2-D electrophoresis
Fill with solution to about 1 cm below the top of the
rectangular glass plate. This height allows 4 to 5 mm
of space for the IPG Strip and an agarose seal). Over-
lay the separating gel as described in step 3.
3
If casting polyacrylamide gels, overlay each gel with
a thin layer of water-saturated n-butanol, water, or
diluted gel buffer to prevent gel exposure to oxygen.
Apply the same amount to each gel to keep the gel
levels the same. Deliver the overlay solution slowly
near the spacer at the side of the sandwich. Allow the
solution to flow across the surface unaided.
4
Allow the gel to polymerize for a minimum of 1 hour.
See page 12 for stack disassembly.
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p7
Note: Avoid repeated and
prolonged contact of
n-butanol with the acrylic
casting chamber to prevent the
acrylic from becoming opaque.