In free flow electrophoresis (FFE) a thin stream of sample is continuously introduced into a planar flow chamber. An electric field is applied perpendicularly to the flow through the separation chamber. Analytes are deflected laterally in the electric field according to their electrophoretic mobility giving rise to individual stream paths. FFE has recently been miniaturized into a microfluidic format (uFFE), requiring less sample and reagents, a simplified flow profile and better heat dissipation.
The continuous nature of uFFE separations suggests a number of novel analytical applications. For example, we have demonstrated how introducing a buffer gradient into the uFFE device can be used to efficiently optimize a range of separation conditions or estimate dissociation constants in as little as five minutes. We have also explored introducing a gradient at the sample channel to titrate a fluorescently labeled aptamer with increasing concentrations of its protein target. Due to the continuous nature of gradient uFFE, complete coverage of the binding curve is possible in as little as five minutes. uFFE is an ideal second stage for two dimensional separations. Continuous uFFE separation allows complete sampling of peaks as they migrate off the first dimension column with minimal loss of efficiency. Coupling to uFFE results in a 10-20 fold improvement in peak capacity without sacrificing separation time. uFFE also offers the potential for impressive microscale purification. We will demonstrate isolation of high affinity aptamers using uFFE.
See more of this Group/Topical: Topical 3: 2011 Annual Meeting of the American Electrophoresis Society (AES)