( lfate TEMED and riboflavin ...)Acrylamide Polymerization A Practical Approach

lfate, TEMED, and riboflavin, are effective. Riboflavin is the better choice for polymerization at low pH (Shi and Jackowski 1998); however, at low pH, TEMED may become protonated. This can result in slower polymerization, since the free base form of TEMED is required for initiation. For acidic buffer systems, alternative initiator systems are sometimes used (Andrews 1990).

Alternative Crosslinkers
PDA (piperazine di-acrylamide), a crosslinking agent that can be substituted for bis in polyacrylamide gels, offers several advantages for electrophoresis. These include reduced background for silver staining, increased gel strength, and higher-resolution gels. PDA can be substituted for bis on a weight basis without changing polymerization protocols.

Crosslinkers other than bis and PDA may be used for specialized purposes, the most common of which is gel solubilization during post-electrophoresis recovery of proteins or nucleic acids. These crosslinkers include DATD (diallyltartardiamide), DHEBA (dihydroxyethylene-bis-acrylamide), and BAC (bis-acrylylcystamine). Alternative crosslinkers may be more or less reactive in polymerization than bis. Therefore, some adjustment in the concentration of initiators may be necessary to achieve optimal polymerization. For a discussion of alternative crosslinkers, see Gelfi and Righetti (1981b).

Gel Additives
The most common gel additives include SDS (sodium dodecyl sulfate), Triton* X-100 detergent, and chaotropic agents such as urea and formamide. Detergents can be added to most common buffer systems without significantly affecting polymerization. Agents such as urea and formamide, however, cause the formation
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