Hoefer SE400 - Manual (Page 44)

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Gel recipes
The Laemmli gel recipes are for 30 ml of a single concentration solution (enough for
one 1.5-mm 18 × 16 cm gel). Tabulated are ingredients and volumes for relatively
large pore gels (7.5 to 10%T range) as well as smaller pore gels (12.5 to 15%T range).
A 4% stacking gel is common. The linear gradient recipe is for 100 ml of solution.
The total volume needed depends on the number of gels cast and the gel thickness;
adjust as necessary. All gels are crosslinked with 2.6%C.
Table 5. Laemmli gel recipes
(per 30 ml resolving gel solution, 5 ml stacking gel solution)
Resolving gel Stacking gel
7.5% 10% 12.5% 15% 4%
Acrylamide stock (Soln. #1) 7.5 ml 10 ml 12.5 ml 15 ml 0.67 ml
1.5 M TrisCl, pH 8.8 (Soln. #2) 7.5 ml 7.5 ml 7.5 ml 7.5 ml
0.5 M TrisCl, pH 6.8 (Soln. #3) 1.25 ml
10% SDS (Soln. #4) 0.3 ml 0.3 ml 0.3 ml 0.3 ml 0.05 ml
Deionized H
2
O 14.6 ml 12.1 ml 9.6 ml 7.1 ml 3.00 ml
10% APS (Soln. #5) 150 µl 150 µl 150 µl 150 µl 25 µl
TEMED 10 µl 10 µl 10 µl 10 µl 2.5 µl
Final Volume 30.0 30.0 ml 30.0 ml 30.0 ml 5.0 ml
For linear gradient gels, use equal volumes of low % and high % acrylamide solutions.
Less APS is added to extend polymerization time, and less still is added to the higher
%T solution to allow polymerization to occur from the top down. In our experience with
the concentrations in the 10–20% gradient example below, ten gel sandwiches can be
poured in a multiple gel caster at a flow rate of 5–10 ml/min.
Table 6. Linear gradient gel recipes (per 100 ml solution)
10% T 20% T
Acrylamide stock (Solution #1) 33.30 ml 66.70 ml
Sucrose — 15.00 g
1.5 M TrisCl, pH 8.8 (Solution #2) 25.00 ml 25.00 ml
10% SDS (Solution #4) 1.00 ml 1.00 ml
Deionized H
2
O to 100.00 ml to 100.00 ml
10% APS (Solution #5) 0.300 ml 0.060 ml
TEMED 0.036 ml 0.036 ml