Cross-linked polyacrylamide coating for capillary isoelectric focusing

Polyacrylamide has been used for capillary wall coating for decades. The coating chemistry includes two main steps: (i) attachment of a bifunctional reagent containing a vinyl group to the silica surface and (ii) extension of the anchored vinyl groups through acrylamide polymerization. Since the introduction of this method, many modifications and improvements have been made. However, few of them are successful for routine capillary isoelectric focusing. One of the major problems is the presence of some microspots in which the silica surfaces are poorly coated. Cross-linking the polyacrylamide molecules anchored around these poorly coated spots seems to be a straightforward solution to this problem. Attempts have been made toward this direction, but cross-linked polyacrylamide coatings have not been demonstrated to be much superior over linear polyacrylamide ones. In this report, we have reexamined this approach and demonstrated that cross-linked polyacrylamide could be excellent for capillary isoelectric focusing. A simple device and a new coating protocol have been developed to produce this coating reliably and reproducibly. Compared to the commercial linear polyacrylamide and hydroxypropyl cellulose coatings for CIEF, the cross-linked polyacrylamide coating is much more stable and robust although their initial performances are comparable.