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We have investigated the interaction of graphene oxide (GO) sheets with supported lipid membranes with focus on how the interaction depends on GO sheet size (three samples in the range of 90-5000 nm) and how it differs between small and large liposomes. The layer-by-layer assembly of these materials into multilamellar structures, as discovered in our previous research, is now further explored. The interaction processes were monitored by two complementary, real time, surface-sensitive analytical techniques: quartz crystal microbalance with dissipation monitoring (QCM-D, electroacoustic sensing) and indirect nanoplasmonic sensing (INPS, optical sensing). The results show that the sizes of each of the two components, graphene oxide and liposomes, are important parameters affecting the resulting multilayer structures. Spontaneous liposome rupture onto graphene oxide is obtained for large lateral dimensions of the graphene oxide sheets.

Aluminum has attracted great attention recently as it has been suggested by several studies to be associated with increased risks for Alzheimer's and Parkinson's disease. The toxicity of the trivalent ion is assumed to derive from structural changes induced in lipid bilayers upon binding, though the mechanism of this process is still not well understood. In the present study we elucidate the effect of Al(3+) on supported lipid bilayers (SLBs) using fluorescence microscopy, the quartz crystal microbalance with dissipation (QCM-D) technique, dual-polarization interferometry (DPI), and molecular dynamics (MD) simulations. Results from these techniques show that binding of Al(3+) to SLBs containing negatively charged and neutral phospholipids induces irreversible changes such as domain formation. The measured variations in SLB thickness, birefringence, and density indicate a phase transition from a disordered to a densely packed ordered phase.

The sequential hybridization of a 534 base pair DNA concatemer layer was monitored by QCM-D and SPR, and the QCM-D data were analyzed by Voigt viscoelastic models. The results show that Voigt-based modeling gives a good description of the experimental data but only if shear viscosity and elasticity are allowed to depend on the shear frequency. The derived layer thickness, shear viscosity and elasticity of the growing film give a representation of the DNA film in agreement with known bulk properties of DNA, and reveal a maximum in film viscosity when the molecules in the layer contain 75 base pairs. The experimental data during construction of a 3084 bp DNA concatemer layer were compared to predictions of the QCM-D response of a 1 μm thick film of rod-like polymers. A predicted nonmonotonous variation of dissipation with frequency (added mass) is in qualitative agreement with the experiments, but with a quantitative disagreement which likely reflects that the flexibility of such long DNA molecules is not included in the model.

Here, the layer-by-layer method was applied to assemble films from chitosan paired with either heparin or a semisynthetic cellulose sulfate (CS) that possessed a higher sulfation degree than heparin. Ion pairing was exploited during multilayer formation at pH 4, while hydrogen bonding is likely to occur at pH 9. Effects of polyanions and pH value during layer formation on multilayers properties were studied by surface plasmon resonance ("dry layer mass"), quartz crystal microbalance with dissipation monitoring ("wet layer mass"), water contact angle, and zeta potential measurements. Bioactivity of multilayers was studied regarding fibronectin adsorption and adhesion/proliferation of C2C12 myoblast cells. Layer growth and dry mass were higher for both polyanions at pH 4 when ion pairing occurred, while it decreased significantly with heparin at pH 9. By contrast, CS as polyanion resulted also in high layer growth and mass at pH 9, indicating a much stronger effect of hydrogen bonding between chitosan and CS. Water contact angle and zeta potential measurements indicated a more separated structure of multilayers from chitosan and heparin at pH 4, while CS led to a more fuzzy intermingled structure at both pH values. Cell behavior was highly dependent on pH during multilayer formation with heparin as polyanion and was closely related to fibronectin adsorption. By contrast, CS and chitosan did not show such dependency on pH value, where adhesion and growth of cells was high. Results of this study show that CS is an attractive candidate for multilayer formation that does not depend so strongly on pH during multilayer formation. In addition, such multilayer system also represents a good substrate for cell interactions despite the rather soft structure. As previous studies have shown specific interaction of CS with growth factors, multilayers from chitosan and CS may be of great interest for different biomedical applications.

The binding of zinc-porphyrin-anchored linear DNA to supported lipid membranes was studied using quartz crystal microbalance with dissipation monitoring (QCM-D). The hydrophobic anchor is positioned at the ninth base of 39-base-pair-long DNA sequences, ensuring that the DNA is positioned parallel to the membrane surface when bound, an important prerequisite for using this type of construct for the creation of two-dimensional (2D) DNA patterns on the surface. The anchor consists of a porphyrin group linked to the DNA via two or three phenylethynylene moieties. Double-stranded DNA where one of the strands was modified with either of these anchors displayed irreversible binding, although binding to the membrane was faster for the derivatives with the short anchor. The binding and subsequent hybridization of single-stranded constructs on the surface was demonstrated at 60 °C, for both anchors, revealing a coverage-dependent behavior. At low coverage, hybridization results in an increase in mass (as measured by QCM-D) by a factor of ~1.5, accompanied by a slight increase in the rigidity of the DNA layer. At high coverage, hybridization expels molecules from the membrane, associated with an initial increase, followed by a decrease in DNA mass (as detected both by QCM-D and by an optical technique). Melting of the DNA on the surface was performed, followed by rehybridization of the single-stranded species left on the surface with their complementary strand, demonstrating the reversibility inherent in using DNA for the formation of membrane-confined nanopatterns., (© 2011 American Chemical Society)

Synthetic lipid bilayers have similar properties as cell membranes and have been shown to be of great use in the development of novel biomimicry devices. In this study, lipid bilayer formation on mesoporous silica of varying pore size, 2, 4, and 6 nm, has been investigated using quartz crystal microbalance with dissipation monitoring (QCM-D), fluorescent recovery after photo bleaching (FRAP), and atomic force microscopy (AFM). The results show that pore-spanning lipid bilayers were successfully formed regardless of pore size. However, the mechanism of the bilayer formation was dependent on the pore size, and lower surface coverages of adsorbed lipid vesicles were required on the surface having the smallest pores. A similar trend was observed for the lateral diffusion coefficient (D) of fluorescently labeled lipid molecules in the membrane, which was lowest on the surface having the smallest pores and increased with the pore size. All of the pore size dependent observations are suggested to be due to the hydrophilicity of the surface, which decreases with increased pore size.

The aim of this work was to create patterned surfaces for localized and specific biochemical recognition. For this purpose, we have developed a protocol for orthogonal and material-selective surface modifications of microfabricated patterned surfaces composed of SiO(2) areas (100 μm diameter) surrounded by Au. The SiO(2) spots were chemically modified by a sequence of reactions (silanization using an amine-terminated silane (APTES), followed by amine coupling of a biotin analogue and biospecific recognition) to achieve efficient immobilization of streptavidin in a functional form. The surrounding Au was rendered inert to protein adsorption by modification by HS(CH(2))(10)CONH(CH(2))(2)(OCH(2)CH(2))(7)OH (thiol-OEG). The surface modification protocol was developed by testing separately homogeneous SiO(2) and Au surfaces, to obtain the two following results: (i) SiO(2) surfaces which allowed the grafting of streptavidin, and subsequent immobilization of biotinylated antibodies, and (ii) Au surfaces showing almost no affinity for the same streptavidin and antibody solutions. The surface interactions were monitored by quartz crystal microbalance with dissipation monitoring (QCM-D), and chemical analyses were performed by polarization modulation-reflexion absorption infrared spectroscopy (PM-RAIRS) and X-ray photoelectron spectroscopy (XPS) to assess the validity of the initial orthogonal assembly of APTES and thiol-OEG. Eventually, microscopy imaging of the modified Au/SiO(2) patterned substrates validated the specific binding of streptavidin on the SiO(2)/APTES areas, as well as the subsequent binding of biotinylated anti-rIgG and further detection of fluorescent rIgG on the functionalized SiO(2) areas. These results demonstrate a successful protocol for the preparation of patterned biofunctional surfaces, based on microfabricated Au/SiO(2) templates and supported by careful surface analysis. The strong immobilization of the biomolecules resulting from the described protocol is advantageous in particular for micropatterned substrates for cell-surface interactions.

The bidirectional transfer of phospholipids between a charged, supported lipid bilayer (SLB) on SiO(2) and oppositely charged, unilamellar vesicles was studied by means of quartz crystal microbalance with dissipation (QCM-D) and optical reflectometry techniques. SLBs and vesicles were prepared from binary mixtures of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) mixed with different fractions of either 1-palmitoyl-2-oleoyl-sn-glycero-3-[phospho-l-serine] (POPS) (negatively charged) or 1-palmitoyl-2-oleoyl-sn-glycero-3-ethylphosphocholine (POEPC) (positively charged). The interaction process consists of an attachment-transfer-detachment (ATD) sequence, where added vesicles first attach to and interact with the SLB, after which they detach, leaving behind a compositionally modified SLB and ditto vesicles. When the process is complete, there is no net addition or reduction of total lipid mass in the SLB, but lipid exchange has occurred. The time scale of the process varies from a few to many tens of minutes depending on the type of charged lipid molecule and the relative concentration of charged lipids in the two membranes. Electrostatically symmetric cases, where only the charge sign (but not the fraction of charged lipid) was reversed between the SLB and the vesicles, produce qualitatively similar but quantitatively different kinetics. The time scale of the interaction varies significantly between the two cases, which is attributed to a combination of the differences in the molecular structure of the lipid headgroup for the positively and the negatively charged lipids used, and to nonsymmetric distribution of charged lipids in the lipid membranes. The maximum amounts of attached vesicles during the ATD process were estimated to be 25-40% of a full monolayer of vesicles, with the precise amount depending on the actual charge fractions in the vesicles and the SLB. Interrupted vesicle exposure experiments, and experiments where the bulk concentration of vesicles was varied, show that vesicles in some cases may be trapped irreversibly on the SLB, when only partial transfer of lipid molecules has occurred. Additional supply of vesicles and further transfer induces detachment, when a sufficient amount of oppositely charged lipids has been transferred to the SLB, so that the latter becomes repulsive to the attached vesicles. Possible mechanistic scenarios, including monomer insertion and hemifusion models, are discussed. The observed phenomena and the actual SLB preparation process form a platform both for studies of various intermembrane molecular transfer processes and for modifying the composition of SLBs in a controlled way, for example, for biosensor and cell culture applications.

Docosahexaenoic acid (DHA) is a polyunsaturated long fatty acid known to have fundamental effects on cell membrane function. Here, the effect of DHA on phosphocholine-supported lipid bilayers was measured using the quartz crystal microbalance with dissipation monitoring (QCM-D) technique. Above a concentration of 60 muM (i.e., near the critical micelle concentration), DHA had drastic effects on the viscoelastic properties of the supported membranes, suggesting a more complex process and structure than simple insertion of molecules in the bilayer. Fluorescence microscopy revealed the spontaneous formation of elongated out-growths from the bilayers, which were remarkable for their length ( approximately 100 mum) and extensive coverage of the surface. These results demonstrate the applicability of QCM-D as a method to screen for conditions where membrane remodeling occurs but also that complementary techniques are required to describe in more detail the changes in viscoelastic properties of the membrane. These results are highly relevant for the present rapid development in the field of model lipid membranes aiming toward increased knowledge about processes occurring at biological surfaces.