Mid-scale free-flow electrophoresis with gravity-induced uniform flow of background buffer in chamber for the separation of cells and proteins

A large-scale free-flow electrophoresis (LS-FFE) is often too large for cell separation of lab scale, whereas micro-FFE (mu FFE) has great difficulty in cell isolation due to easy blockage by cell accumulation in mu FFE. In this study, a mid-scale FFE (MS-FFE) is developed for cell and protein separations. The volume of the separation chamber (70 x 40 x 0.1-0.8 mm) is from 280 mu L to 2.24 mu L, much lower than that in an LS-FFE but higher than that in a mu FFE. Gravity is used for uniform flow of the background buffer only via a single pump with 16 channels and the sample is injected via an adjuster originally used for clinical intravenous injection. The experiments reveal that the hydrodynamic and electro-hydrodynamic flows are much stable, and the Joule heat can be effectively dispersed without obvious positive or negative deviation as shown by the omega plots. By the device, Escherichia coli and Staphylococcus aureus, which easily accumulate to block mFFE and are separated with difficulty due to their same negative charges carried, can be well isolated under the conditions of 4.5 mM pH 8.5 Tris-boric buffer (4.5 mM Tris, 4.5 mM boric acid) with 0.10 mM ethylene diamine tetraacetic acid and 5% m/v sucrose, 200 mu L/min, 800 V, and sample injection via inlet 4. The mid-scale FFE device could also be used for the separation of three model proteins of horse heart cytochrome c, myoglobin and bovine serum albumin. The device has clear significance for mid-scale separation of cells and proteins.