Abstract: Microchip free flow electrophoresis (μFFE) is a significant microscale technique for the continuous pre-fractionation and the preparation of valuable biological samples. In our recent work, monolithic polyacylamide (PAM) materials were polymerized in microchamber by ultraviolet (UV) initiated polymerization. With the further immobilization of a stable pH gradient on the monolith, a novel microchip free flow isoelectric focusing (μFF-IEF) with monolithic immobilized pH gradient (M-IPG) materials was developed, by which fluorescein-5-isothiocyanate (FITC) labeled glycin, proline and lysine, with a minimum pI difference of 0.33 units, were well separated with a resolution higher than that performed by traditional μFF-IEF. Our experimental results demonstrate that by μFF-IEF with M-IPG, not only the interference of mobile carrier ampholytes in buffer, usually indispensable in traditional μFF-IEF, on the further separation by other techniques and the identification by mass spectrometry (MS) could be avoided, but also the improved resolution and detection sensitivity could be obtained compared with traditional μFF-IEF. Therefore, such a novel technique might be promising in microscale consecutive separation and preparation of samples.

Key words: amino acids , immobilized pH gradient, isoelectric focusing, monolithic materials, microchip free flow electrophoresis