Electropreconcentration, gate injection, and capillary electrophoresis separation on a microchip

The nanochannel-based electropreconcentration is not compatible with successive capillary zone electrophoresis (CZE). In this study, the incompatibility is theoretically discussed and experimentally proven, and then, the development of a monolithic glass microfluidic chip for performing integrated electropreconcentration and CZE separation is described. The sample is electropreconcentrated at the interface of a micro- and nanochannel where electric double layer overlap conditions exist. Because an ion-depletion region develops at the leading front of the preconcentrated plug, a field-enhanced sample stacking effect occurs which limits the separation efficiency unless compensated for. The ion-depletion region was confirmed by monitoring the solution conductivity at discrete points in the microchannel during the preconcentration step. The solution conductivity decreased >20-fold during the preconcentration step. To overcome the effects of this region, a cross-intersection was used to shunt the ion-depleted buffer away from the analysis channel while reintroducing the running buffer. When the preconcentrated sample plug arrives at the cross-intersection, it is gate injected into the analysis channel so that fresh running buffer surrounds the plug. Under these conditions, three-peptide mixture was preconcentrated ∼200-fold in 60 s and the preconcentrated plug was successfully resolved with better than 1% relative standard deviations in migration times.