Mini-review Received: 14 April 2014 Revised: 23 June 2014 Accepted article published: 27 June 2014 Published online in Wiley Online Library: (wileyonlinelibrary.com) DOI 10.1002/jctb.4473 Membranes for separation of biomacromolecules and bioparticles via ﬂow ﬁeld-ﬂow fractionation Ulku Bade Kavurt, a Maria Marioli, b Wim Th. Kok b and Dimitrios Stamatialis a Abstract Flow ﬁeld-ﬂow fractionation (FlFFF) is a liquid-phase separation technique that can separate macromolecules and particles on the basis of diﬀerences in their diﬀusion coeﬃcient, and therefore their size. The membrane in the FlFFF channel is one of the most important elements involved in accomplishing the separation. This mini-review focuses on requirements and important points related to membranes used in FlFFF channels. Unless it is selected properly, the membrane can be a weak point of the technique due to risks such as sample loss because of the interactions between the sample and the membrane. © 2014 Society of Chemical Industry Keywords: membrane; ﬂow ﬁeld-ﬂow fractionation; biomacromolecules; bioparticles INTRODUCTION Field-ﬂow fractionation (FFF) is a class of analytical techniques that aims to separate macromolecules, supramolecules and micron-sized particles. The unique characteristic of FFF is the ability to separate an extraordinary range of molecular weight substances without loss of resolution. It is applicable to analytes of molecular weight from 10 3 g mol –1 to 10 18 g mol –1 , which corresponds to a size of a few nanometers to many micrometers, respectively. FFF was introduced in 1966 by J. Calvin Giddings 1 who was not only the pioneer but he also formulated a rigorous theoretical basis that paved the way for further developments. 2,3 Unlike chromatography, FFF lacks a stationary phase and the separation mechanism is based solely on the non-uniformity of a convective ﬂow and a lateral concentration gradient caused by an external ﬁeld. Recently, a critical review of FFF in general 4 and reviews with speciﬁc focus on biomolecules and bioparticles, 5 – 7 polymers, 8 food macromolecules 9 and natural colloids 10 have been published. With respect to the nature of the applied ﬁeld, FFF is classiﬁed as electrical ﬁeld-ﬂow fractionation (ElFFF), thermal ﬁeld-ﬂow frac- tionation (ThFFF), sedimentation ﬁeld-ﬂow fractionation (SdFFF), ﬂow ﬁeld-ﬂow fractionation (FlFFF), gravitational ﬁeld-ﬂow fractionation GrFFF), dielectrophoretic ﬁeld-ﬂow fractionation (DEP-FFF), magnetic ﬁeld-ﬂow fractionation (MgFFF) and isoelec- tric focusing ﬁeld-ﬂow fractionation (IEFFFF). Among them, FlFFF is especially intriguing for bioanalysis as it operates under mild conditions enabling the characterization of intact biomolecules and bioparticles. It has been used extensively for the separation of proteins, polysaccharides, nucleic acids, lipoproteins, liposomes, viruses, cell organs and cells. 7,11 The membrane used in the FlFFF channel is considered as its heart. 12 This mini-review focuses on requirements and important points related to membranes used in FlFFF channels for bioanaly- sis. Here, we mostly review studies on biomacromolecules, lipopro- teins, virus-like particles, and liposomes, while for environmental applications of FlFFF such as characterization of natural colloids in water, the reader is referred to other relevant reviews. 10,13,14 INTRODUCTION-FIELD-FLOW FRACTIONATION (FFF) The separation device, in the majority of the subtechniques that constitute the FFF family, is a ribbon-like channel with typically a thickness of 50–500 μm, a width of 0.5–3 cm and a length of 10–50 cm. This thin geometry provides the low Reynolds num- ber required for laminar longitudinal ﬂow (parabolic ﬂow proﬁle), while the rectangular cross-section of high aspect ratio increases the mass load capacity and minimizes the edge eﬀects. An exter- nal ﬁeld imposed transversely to the longitudinal ﬂow and along the thinnest dimension (here arbitrarily assigned as the x-axis), causes migration of the analytes towards one wall of the chan- nel, the so-called accumulation wall (Fig. 1(a)). Under the com- bined action of lateral diﬀusion and drift velocity the system reaches quasi-equilibrium. At this time, the concentration has an exponential proﬁle which can be described mathematically as C(x) = C 0 exp(-x/l), where l is the mean thickness of the accu- mulation layer determined by the interaction of the molecules with the applied ﬁeld. 2 Consequently, sample components with ∗ Correspondence to: Dimitrios Stamatialis, Department of Biomaterials Sci- ence and Technology, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, P.O.Box 217, 7500 AE Enschede, The Nether- lands. E-email: d.stamatialis@utwente.nl a Department of Biomaterials Science and Technology, MIRA Institute for Biomedical, Technology and Technical Medicine, University of Twente, P.O.Box 217, 7500, AE Enschede, The Netherlands b Analytical Chemistry Group, van’t Hoﬀ Institute for Molecular Sciences, Univer- sity of Amsterdam, Postbus 94157, 1090 GD Amsterdam, The Netherlands J Chem Technol Biotechnol (2014) www.soci.org © 2014 Society of Chemical Industry