Bio-Rad Model 111 Mini IEF Cell - Manual (Page 12)

3.5 Casting Polyacrylamide Gels
Important: The Glass Plates and the casting tray must be clean and dry. Do not use organic
solvents, abrasive cleaners, or hot water on the casting tray. Clean the Glass Plates with soap
and water and rinse with deionized water followed by ethanol. Wipe the plates dry with lint-
free paper.
1. With the gel support film facing down, place the Glass Plate(s) on the casting tray so that it
rests on the spacer bars.
2. Prepare the monomer-ampholyte solution (see Section 3.3). Degas the solution for 5 minutes
under vacuum. Do not degas longer, as a slight amount of O
2
is required to catalyze
riboflavin-5-phosphate.
4. Prepare the catalyst solutions.
Note: Always use freshly prepared persulfate solutions
5. Add the catalyst solutions to the degassed monomer and swirl gently.
Caution: Do not mouth pipet acrylamide solutions. Wear gloves. Acrylamide is a neurotoxin!
6. Pipet the solution between the glass plate and the casting tray as shown in Figure 3.1.
a. Hold the pipet at a 45° angle and clear the air bubble from the tip.
b. Start the monomer flow at one end of the glass plate and slowly move the pipet to the
other spacer.
c. When a liquid front is established across the plate, slowly add the remaining monomer
from the midpoint of the plate.
d. Control the flow rate to prevent air bubbles. If a bubble is trapped, remove it by sliding the
plate sideways until the bubble is at the edge.
Fig. 3.1. Injecting monomer/ampholyte solution in casting tray.
8
a. Clear bubble from end of pipet. b. Start solution at one corner of
plate and move it along the edge to form a front. Be sure the
solution contacts both the plate and tray. Avoid trapping air
bubbles by releasing the solution slowly.
c. Continue to inject solution, keeping the
front fairly even to avoid forming bubbles.
b a
Remove
this bubble