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3.10 Perpendicular Gradient Formation-continued
10. Remove comb and quickly transfer gel sandwich to cassette by removing all #2 clamps.
LEAVE GEL WRAP IN PLACE
. The Gel Wrap acts as a barrier and prevents perpendicular
electrical fields from intenferring with outside lanes. Re-clamp to cassette using #1 clamps/4
per side. Dislodge the bottom section of the Gel Wrap from the plates to allow buffer contact
with the gel. See
figure 3.8
(page 18
).
11. Immerse cassette into pre-heated buffer tank, and attach recirculating tubes to each
upper reservoir (
fig. 3.8, 3.9a,b,c).
Rinse loading interface of gel with tank buffer to remove
non-polymerized acrylamide or excess urea. Rinse each well with buffer and fill upper
reservoir
12. Load samples.
13. Turn on power supply to 150V (40mA) constant Volts for 5-7 hours.
14. When run is completed, turn off power supply, disconnect recirculating tubes and power
leads, remove cassettes, remove glass sandwiches from cassettes.
15. Using a wedge plate separator,cat. # WPS-100, pry plates apart and immerse gel/plate in
buffer tray. Stain with EtBr (1-5ug/ml) for 5-10 minutes. Lift floating gel on plate out of tray and
rinse with DD H 0. Flip over onto saran wrap and peal off glass plate. View bands on
transilluminator. Electro-Blot or photograph.
