Focusing of Proteins in a Horseshoe Microchannel

Ampholyte based isoelectric focusing (IEF) simulation was conducted to study dispersion of proteins in a horseshoe microchannel. Four model proteins (pls = 6.49, 7.1, 7.93 and 8.6) are focused in a 1 cm long horseshoe channel under an electric field of 300 V/cm. The pH gradient is formed in the presence of 25 biprotic carrier ampholytes (ΔpK = 3.0) within a pH range of 6 to 9. The proteins are focused at 380 sec in a nominal electric field of 300 V/cm. Our numerical results show that the band dispersions of a protein are large during the marching stage, but the dispersions are significantly reduced when the double peaks start to merge. This rearrangement of spreading band is very unique compared to linear electrokinetic phenomena (capillary electrophoresis, zone electrophoresis or electroosmosis) and is independent of channel position and channel shape. Hence, one can perform IEF in complex geometries without incorporating hyperturns.

Microfluidic Devices for Rapid Protein Separation

We present our investigation of using microfluidic devices for rapid protein separation. The devices were made from cyclic olefin copolymers that have high optical clarity and high glass transition temperature. Protein separation was achieved by using isoelectric focusing (IEF) and polyacrylamide gel electrophoresis (PAGE). A laser-induced fluorescence (LIF) imaging system was developed to detect proteins while they migrated under an electric field. IEF was carried out in a separation medium consisting of carrier ampholytes and a mixture of linear polymers. Dynamic coating of the linear polymers prevented proteins from adsorption and suppressed electroosmotic flows. PAGE was achieved in twenty-nine parallel channels. In addition, we integrated IEF with PAGE in a microfluidic device for two-dimensional protein separation.