Capillary isoelectric focusing with an open tubular immobilized pH gradient.

Capillary isoelectric focusing (CIEF) was first performed in an open tubular mode. It depended on an immobilized pH gradients from the polymerization of several solutions, which contained acrylamide, N,N'-methylenebisacrylamide, allylamine, and acrylic acid. Different ratios of the basic and acidic monomers lead to solutions of various pHs. When injected into a capillary in the sequence of their pHs, the solutions polymerized and resulted into polymers with a series of isoelectric points (pIs). These polymers formed a covalently attached hydrophilic layer in the capillary, acting as both column bonding and carrier ampholytes. When five polymers were orderly lined up in a capillary, an immobilized pH gradient was established. This open tubular column was subjected to CIEF, showing excellent performance properties for protein focusing.