( Swirl the solutio...)Acrylamide Polymerization A Practical Approach

Swirl the solution gently but thoroughly.

Cast the gel and insert the well-forming comb without trapping air under the teeth.

Allow polymerization to occur at room temperature at least 90 min prior to use (see Figure 1).

Photochemical Polymerization
This protocol is recommended for isoelectric focusing (IEF) gels. Since molecules remain in the IEF gel during electrophoresis, excess ions from initiators can cause distortion of bands. Photochemical initiation is recommended for IEF gels because it is effective at low initiator concentrations.

1. When the gel solution is prepared and brought to room temperature (2325C), degas the solution under a vacuum of 125 torr or better for 2 min at room temperature (for systems in which constant agitation is used for degassing, 1 min is sufficient).

2. Add initiators as follows:

3. Swirl the solution gently but thoroughly.

4. Cast the gel and allow polymerization to occur for at least 2 hr for isoelectric focusing gels. If separation is to be based on size, allow photochemically initiated gels, to polymerize for 8 hr under light from a nearby fluorescent lamp, (Righetti et al. 1981).

Polymerization Analysis
There are several ways to assess the extent and reproducibility of polymerization. One of the easiest methods is to routinely monitor the time required for visible gelation. There are several factors which affect the polymerization rate. A significant change in the time required for visible gelation indicates that one of the parameters has changed.

The polymerized gel should be inspected for evidence of inhibition or nonuniform polymerizati
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