Miniaturizing free‐flow electrophoresis – a critical review

Abstract

Free-flow electrophoresis (FFE) separation methods have been developed and investigated for around 50 years and have been applied not only to many types of analytes for various biomedical applications, but also for the separation of inorganic and organic substances. Its continuous sample preparation and mild separation conditions make it also interesting for online monitoring and detection applications. Since 1994 several microfluidic, miniaturized FFE devices were developed and experimentally characterized. In contrast to their large-scale counterparts microfluidic FFE (mu-FFE) devices offer new possibilities due to the very rapid separations within several seconds or below and the requirement for sample volumes in the microliter range. Eventually, these mu-FFE systems might find application in so-called lab-on-a-chip devices for real-time monitoring and separation applications. This review gives detailed information on the results so far published on mu-FFE chips, comprising its four main modes, namely free-flow zone electrophoresis (FFZE), free-flow IEF (FFIEF), free-flow ITP (FFITP), and free-flow field-step electrophoresis (FFFSE). The principles of the different FFE modes and the basic underlying theory are given and discussed with special emphasis on miniaturization. Different designs as well as fabrication methods and applied materials are discussed and evaluated. Furthermore, the separation results shown indicate that similar separation quality with respect to conventional FFE systems, as defined by the resolution and peak capacity, can be achieved with mu-FFE separations when applying much lower electrical voltages. Furthermore, innovations still occur and several approaches for hyphenated, more integrated systems have been proposed so far, some of which are discussed here. This review is intended as an introduction and early compendium for research and development within this field.

Proteomics Methodology Applied to the Analysis of Filamentous Fungi - New Trends for an Impressive Diverse Group of Organisms

“…In all studies, proteins primarily involved in translation, energy metabolism, transport processes and the stress response were most abundant. Penicillium chrysogenum, the microorganism industrially used for the production of penicillin, is another example of filamentous fungus whose genome sequence has been recently released (van den Berg et al, 2008). In this specific case, a first study was reported about the intracellular proteome reference map (Jami et al, 2010a).…”

“…In spite of its popularity, the time consumption (2-3 hours) of the process and batch enzymes variability are cons of this method. Secondly, different methods can be used for debris elimination as filtration; low-speed centrifugation; differential detergent fractionation; centrifugal elutriation; ultracentrifugation (linear or density gradients); immunomagnetic separation, which has been used for organelle isolation (mitochondria vacuoles, microbodies, endosomes, vesicles) in other eukaryotes; or miniaturized free-flow electrophoresis (FFE) (Kohlheyer et al 2008, Ferreira de Oliveira & de Graaff, 2011. A large number of diseases and developmental abnormalities result from mitochondrial functions.…”

Experimental and Simulation Research of the Separation of Amino Acids on Micro Free-Flow Electrophoresis Chip with Voltage Applied in Two-Dimensions

“…Compared with other separation techniques on a microchip, it has shown great advantages as a continuous separation method, which requires less buffers and samples, simplifies the liquid manifold and improves the impact of Joule heat on electrophoresis. 3 Many kinds of modes can be used on the μFFE chip. 4 Among them, free-flow zone electrophoresis and free-flow isoelectric focusing have recently gained much attention and progress.…”

A PDMS Sample Pretreatment Device for the Optimization of Electrokinetic Manipulations of Blood Serum

“…Free-flow electrophoresis (FFE) is a common separation technique that has been in use since the 1960's and applied to microfluidic since 1994 [37]. Like CE, FFE uses an electric field to aid in the separation of solutes from each other in an electrolytic buffer solution.…”

“…This method of FFE uses buffers with a constant pH and conductivity to separate different solutes based on their mobility, which is determined by their size-to-charge ratio. Even though CE is primarily used for analysis of analytes and FFZE is used for analyte separations, there is a direct correlation between the operation of the two [37]. For information concerning the comparison of the underlying theory of both techniques, please refer to Kasicka's paper on this topic [38].…”