Non-Polar Electrical Field Flow Fractionation (Np-EFFF)
We report the application of non-polar Electrical Field-Flow Fractionation (Np-EFFF) to the isolation and characterization of membrane bound proteins. These proteins are presently understudied, despite the fact that about 25-30% of the human genome encodes for them. Present methods, e.g. electrophoresis, denature lipophilic and membrane proteins by adding surfactants to suspend them in aqueous media. They also employ high applied voltages causing thermal degradation, and they force fragile proteins through packed gel beds causing mechanical damage. The Np-EFFF method consists of an ultra thin, open channel defined by two oppositely charged conductive plates. A non-polar medium is passed through the channel in laminar flow profile, and the protein sample is injected into the fluid. As the sample passes between the electrically charged plates, proteins separate into different regions of the flow profiles based on their electrophoretic mobility and size. Theory predicts and previous results show that surface roughness of the conductive plates is a significant degradation to the resolving power of the method. We use astronomy-quality gold mirrors with surface roughness less than 1/10 λ. On-axis fittings replace formerly angled fittings to minimize turbulence at channel entrance and exit. The aim of these engineering changes is to better separate the proteins. Collected fractions may then be characterized for enzyme activity.
Walker, Samantha, "Non-Polar Electrical Field Flow Fractionation (Np-EFFF)" (2015). College of Arts and Sciences Presentations. Paper 14.