Search

A novel setup was recently developed, combining quartz crystal microbalance with dissipation monitoring (QCM-D) and optical reflectometry for measurements on one and the same surface of, for example, biomolecular adlayers and interactions (Wang et al. Rev. Sci. Instr. 2008, 79 075107). This combination was chosen on the basis of prior experience of using QCM-D and optical techniques in separate instruments, which showed both the advantage of employing multiple techniques and the disadvantage of not working with the same surface and (flow) cell. The new instrument provides, for example, information about associated water and structural changes of the adlayers that would often pass unnoticed or be hard to interpret or quantify, using either technique alone. The triple response instrument (QCM-D frequency and dissipation and reflectometry) is here applied to four model systems: (A) formation of supported lipid bilayers (SLBs), (B) lipid exchange between a SLB and transiently adsorbed vesicles, (C) binding of a hydrated peptide on a functionalized SLB, and (D) streptavidin coupling to a biotinylated SLB, followed by attachment of biotinylated vesicles. The results demonstrate three major advantages of the combination instrument: (i) much faster data collection because the experiments are done on one surface for all signals, (ii) a common time axis and the same relative importance of surface kinetics and mass transport because the same liquid sample and the same transport conditions apply, and (iii) new features are discovered about the studied system that would be difficult to unravel in separate instruments.

The thermal stability of hydroxyalkanethiolate self-assessed monolayers (SAM) on gold can be improved by introducing an amide group in the vicinity of the thermal group. The study reveals that lateral hydrogen bonding, introduced via amide groups, is a convenient way to improve the thermal stability of alkanthiolate SAMs.

Research has been conducted on the oligo(ethylene glycol)-terminated alkanethiol amides. The synthesis of the these amides and analogous esters has been conducted via attaching hydrophobic tails at the far end of the hexa(ethylene glycol) chains and the results are discussed.

Plasminogen is a precursor to the fibrinolytic enzyme plasmin and is known to undergo large conformational changes when subjected to low molecular lysine analogues such as tranexamic acid (TA) or [epsilon]-amino-n-caproic acid (EACA). Here, we demonstrate how well-controlled surface immobilization of biotinylated plasminogen allows for monitoring of the interaction between TA and EACA with plasminogen. The interaction was studied by the quartz crystal microbalance with dissipation monitoring (QCM-D) technique as well as by surface plasmon resonance (SPR) based sensing. QCM-D measures changes in acoustically coupled mass (by detection of changes in the resonance frequency of the crystal, [DELTA]f) and is sensitive to changes in mass adsorbed on the sensor surface including how liquid medium is associated with this material. Through the dissipation factor (i.e., changes in the energy dissipation of the crystal oscillation, [DELTA]D), QCM-D is also sensitive to the viscoelastic properties of material adsorbed to the sensor surface. Upon binding of TA or EACA, changes in the plasminogen structure were recorded as distinct, although small, [DELTA]D responses which were used to determine affinity constants. By comparing native and truncated plasminogen, we conclude that the observed dissipation shifts were caused by conformational changes in the proteins leading to changes in the viscoelastic properties of the protein layer on the surface. These results demonstrate a novel application of the QCM-D technique, paving the way for a whole new approach to screening of this target for novel lead structures. 10.1021/ac1016419

An example is provided on how lipid transfer is used to form lipid layers with striking new features with respect to stability. The results have provided evidence for lipid transfer between the initial POEPC supported lipid bilayer (SLB) and D31-POPS vesicles in solution and the reassembled SLB has a major asymmetry between the two leaflets.

A comparative study of the self-assembly and phase behavior of seven different oligo(ethylene glycol) (OEG)-terminated alkanethiols on polycrystalline gold surfaces is presented. The results from contact angle goniometry, null ellipsometry, and infrared reflection, absorption spectroscopy (IRAS) indicate that the intermolecular interactions can be partly used to control the conformation and order of the OEG portion of the self-assembled monolayers (SAMs).

The phase behavior of oligo(ethylene glycol) self assembled monolayers using temperature programmed infrared reflection adsorption spectroscopy are explored. The oligomer conformation is marginally affected upon increasing the temperature to 75 degree Celsius, displaying the dominating all-trans phase.

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. [ABSTRACT FROM AUTHOR]

There is an urgent need for improved models for cardiotoxicity testing. Here we propose acoustic sensing applied to beating human cardiomyocyte clusters for non-invasive, surrogate measuring of the QT interval and other characteristics of the contractile machinery. In experiments with the acoustic method quartz crystal microbalance with dissipation monitoring (QCM-D), the shape of the recorded signals was very similar to the extracellular field potential detected in electrochemical experiments, and the expected changes of the QT interval in response to addition of conventional drugs (E-4031 or nifedipine) were observed. Additionally, changes in the dissipation signal upon addition of cytochalasin D were in good agreement with the known, corresponding shortening of the contraction-relaxation time. These findings suggest that QCM-D has great potential as a tool for cardiotoxicological screening, where effects of compounds on the cardiomyocyte contractile machinery can be detected independently of whether the extracellular field potential is altered or not. [ABSTRACT FROM AUTHOR]

The sequential hybridization of a534 base pair DNA concatemerlayer was monitored by QCM-D and SPR, and the QCM-D data were analyzedby Voigt viscoelastic models. The results show that Voigt-based modelinggives a good description of the experimental data but only if shearviscosity and elasticity are allowed to depend on the shear frequency.The derived layer thickness, shear viscosity and elasticity of thegrowing film give a representation of the DNA film in agreement withknown bulk properties of DNA, and reveal a maximum in film viscositywhen the molecules in the layer contain 75 base pairs. The experimentaldata during construction of a 3084 bp DNA concatemer layer were comparedto predictions of the QCM-D response of a 1 μm thick film ofrod-like polymers. A predicted nonmonotonous variation of dissipationwith frequency (added mass) is in qualitative agreement with the experiments,but with a quantitative disagreement which likely reflects that theflexibility of such long DNA molecules is not included in the model. [ABSTRACT FROM AUTHOR]

Supported lipid bilayers (SLBs) at surfaces provide a route to quantitatively study molecular interactions with and at lipid membranes via different surface-based analytical techniques. Here, a method to fabricate SLBs with controlled curvatures, in the nanometer regime over large areas, is presented, utilizing lipid vesicle rupture onto nanostructured sensor substrates. Heat treated colloidal particle masks were used as templates to produce silicon dioxide films with systematically varied radius of curvature (ROC, 70 to 170 nm are demonstrated) and quartz crystal microbalance with dissipation monitoring (QCM-D) was used to confirm vesicle rupture onto such structured surfaces. Fluorescence microscopy was used to show fluidity of the supported membranes. The formation of confluent SLBs is demonstrated at the nanostructured surfaces from vesicles composed of POPC lipids. However, at surfaces with decreasing ROCs, vesicle rupture was hindered but with an increasing fraction of the positively charged lipid POEPC in the vesicles, it was possible to form good quality supported bilayers on all curvatures studied. Curved SLBs open up the possibility to systematically study the influence of curvature on molecular interactions at lipid membranes. [ABSTRACT FROM PUBLISHER]

We have developed and characterized novel biomimetic membranes, formed at nanostructured sensor substrates with controlled curvatures, motivated by the many biological processes that involve membrane curvature. Model systems with convex nanostructures, with radii of curvatures (ROCs) of 70, 75, and 95 nm, were fabricated utilizing colloidal assembly and used as substrates for supported lipid bilayers (SLBs). The SLBs were formed via vesicle adsorption and rupture, and the vesicle deposition pathway was studied by means of quartz crystal microbalance with dissipation (QCM-D) and fluorescence microscopy. SLBs conforming to the underlying nanostructured surfaces, which exhibit increased surface area with decreased ROC, were confirmed from excess mass, monitored by QCM-D, and excess total fluorescence intensities. The formation of SLBs at the nanostructured surfaces was possible, however, depending on the ROC of the structures and the lipid vesicle charge the quality varied. The presence of nanostructures was shown to impair vesicle rupture and SLB formation was progressively hindered at surfaces with structures of decreasing ROCs. The introduction of a fraction of the positively charged lipid POEPC in the lipid vesicle membrane allowed for good quality and conformal bilayers at all surfaces. Alternatively, for vesicles formed from lipid mixtures with a fraction of the negatively charged lipid POPS, SLB formation was not at all possible at surfaces with the lowest ROC. Interestingly, the vesicle adsorption rate and the SLB formation were faster at surfaces with nanostructures of progressively smaller ROCs at high ratios of POPS in the vesicles. Development of templated SLBs with controlled curvatures provides a new experimental platform, especially at the nanoscale, at which membrane events such as lipid sorting, phase separation, and protein binding can be studied. [ABSTRACT FROM AUTHOR]

A novel set-up combining the quartz crystal microbalance with dissipation monitoring technique (QCM-D) and electrochemical impedance spectroscopy (EIS) under flow conditions was successfully used to follow supported lipid bilayer (SLB) formation on SiO2. This study demonstrates the simultaneous detection, in real time, of both the electrical and the structural properties of the SLB. The combination of the two techniques provided novel insights regarding the mechanism of SLB formation: we found indications for an annealing process of the lipid alkyl chains after the mass corresponding to complete bilayer coverage had been deposited. Moreover, the interaction of the SLB with the pore-forming toxin, gramicidin D (grD) was studied for grD concentrations ranging from 0.05 to 40 mg L−1. Membrane properties were altered depending on the toxin concentration. For low grD concentrations, the electrical properties of the SLB changed upon insertion of active ion channels. For higher concentrations, the QCM-D data showed dramatic changes in the viscoelastic properties of the membrane while the EIS spectra did not change. AFM confirmed significant structural changes of the membrane at higher grD concentrations. Thus, the application of combined QCM-D and EIS detection provides complementary information about the system under study. This information will be particularly important for the continued detailed investigation of interactions at model membrane surfaces. [ABSTRACT FROM AUTHOR]

The importance of the lipid phase on the formation of supported lipid bilayers (SLBs) viavesicle fusion and on the resulting SLB homogeneity at SiO2surfaces has been studied by the quartz crystal microbalance with dissipation (QCM-D) monitoring technique. Physiologically relevant lipid compositions were chosen to correspond to different regions (ld, loand coexistence of phases) in established phase diagrams of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), N-palmitoyl-d-erythro-sphingosylphosphorylcholine (PSM) and cholesterol. For most compositions, SLBs formed through vesicle rupture in a critical-surface-coverage dependent manner. Inclusion of PSM and cholesterol into POPC vesicles significantly impaired the vesicle rupture process such that a higher critical concentration of vesicles on the surface was needed before the rupture process started. When increasing the cholesterol content the vesicles formed SLBs containing more defects in the form of intact vesicles adsorbed on the surface up to a point (lophase) where vesicles did not break at all but formed supported vesicular layers. The hampering of vesicle rupture is interpreted in terms of the ability of cholesterol to accommodate vesicle deformation. Experiments using elevated temperatures to alter the lipid membrane into a more fluid phase significantly improved the quality of the SLB showing the importance of both cholesterol content and the lipid phase on SLB homogeneity. [ABSTRACT FROM AUTHOR]

A novel setup was recently developed, combining quartz crystal microbalance with dissipation monitoring (QCM-D) and optical reflectometry for measurements on one and the same surface of, for example, biomolecular adlayers and interactions (Wang et a!. Rev. Sci. Instr. 2008, 79 075107). This combination was chosen on the basis of prior experience of using QCM-D and optical techniques in separate instruments, which showed both the advantage of employing multiple techniques and the disadvantage of not working with the same surface and (flow) cell. The new instrument provides, for example, information about associated water and structural changes of the adlayers that would often pass unnoticed or be hard to interpret or quantify, using either technique alone. The triple response instrument (QCM-D frequency and dissipation and reflectometry) is here applied to four model systems: (A) formation of supported lipid bilayers (SLBs), (B) lipid exchange between a SLB and transiently adsorbed vesicles, (C) binding of a hydrated peptide on a functionalized SLB, and (D) streptavidin coupling to a biotinylated SLB, followed by attachment of biotinylated vesicles. The results demonstrate three major advantages of the combination instrument: (i) much faster data collection because the experiments are done on one surface for all signals, (ii) a common time axis and the same relative importance of surface kinetics and mass transport because the same liquid sample and the same transport conditions apply, and (iii) new features are discovered about the studied system that would be difficult to unravel in separate instruments. [ABSTRACT FROM AUTHOR]

We describe a novel method to immobilize recombinant human tissue factor (rhTF) reconstituted in phospholipid vesicles. The rhTF vesicles were immobilized in a multilayer vesicle structure using cholesterol–DNA tethers spontaneously inserted into the lipid membrane. The properties of the rhTF vesicle surface modification were characterized by surface plasmon resonance biosensor technology. As an application of this surface modification, we investigated its use as a blood coagulation activating surface. The coagulation activating capacity of the surface modification was tested by exposure to human whole blood in a flow chamber. No increase in rhTF levels in the blood was found after passage through the flow chamber, indicating that the rhTF surface modification was stable. Thrombin–antithrombin (TAT) and prothrombin fragment (PF) 1 + 2 levels increased after exposure to the surface, and decreased in a concentration-dependent way upon addition of melagatran (a direct thrombin inhibitor), i.e., coagulation activity triggered by rhTF could be suppressed by anticoagulation. The results with this new thrombogenic surface are promising, and will be further developed into a useful tool for coagulation related investigations, e.g., characterization of anticoagulants and biomaterials. [ABSTRACT FROM AUTHOR]

We have developed an instrument for surface interaction studies, which combines a newly invented four detector optical reflectometry setup with quartz crystal microbalance with dissipation (QCM-D) monitoring. The design is such that data from both techniques can be obtained simultaneously on the same sensor surface, with the same signal-to-noise ratio and time resolution, as for the individual techniques. In addition, synchronized information about structural transformations, molecular mass, and the hydration of thin films on solid surfaces can be obtained on the same specimen, as validated by monitoring the formation of supported lipid bilayers on a silica-coated QCM sensor surface. We emphasize that the optical (molecular) mass can be separated from the acoustic mass including hydrodynamically coupled solvent, which means, in turn, that the amount of solvent sensed by the QCM-D technique can be dynamically resolved during adsorption processes. In addition, the advantage/necessity to use four, compared to two, detector reflectometry is emphasized. [ABSTRACT FROM AUTHOR]

Low protein adsorption is believed advantageous for blood-contacting materials and ethylene glycols (EG)-based polymeric compounds are often attached to surfaces for this purpose. In the present study, the adsorption of fibrinogen, serum, and plasma were studied by ellipsometry on a series of well-defined oligo(EG) terminated alkane-thiols self-assembled on gold. The layers were prepared with compounds of the general structure HS-(CH[sub 2])[sub 15]-CONH-EG[sub n], where n = 2, 4, and 6. Methoxy-terminated tri(EG) undecanethiol and hydroxyl-terminated hexadecanethiol self-assembled monolayers (SAMs) were used as references. The results clearly demonstrate that the adsorption depends on the experimental conditions with small amounts of fibrinogen adsorbing from a single protein solution, but larger amounts of proteins from serum and plasma. The adsorption of fibrinogen and blood plasma decreased with an increasing number of EG repeats and was temperature-dependent. Significantly less serum adsorbed to methoxy tri(EG) than to hexa(EG) and more proteins remained on the latter surface after incubation in a sodium dodecyl sulfate (SDS) solution, indicating a looser protein binding to the methoxy-terminated surface. All surfaces adsorbed complement factor 3 (C3) from serum and plasma, although no surfacemediated complement activation was observed. The present study points to the importance of a careful choice of the protein model system before general statements regarding the protein repellant properties of potential surfaces can be made. [ABSTRACT FROM AUTHOR]

This article summarizes our most recent contributions to the rapidly growing field of supported lipid assemblies with emphasis on current studies addressing both fundamental and applied aspects of supported lipid bilayer (SLB) and tethered lipid vesicles (TLVs) to be utilized in sensing applications. The new insights obtained from combining the quartz crystal microbalance with dissipation monitoring technique with surface plasmon resonance are described, and we also present recent studies in which nanoplasmonic sensing has been used in studies of SLBs and TLVs. To gain full control over the spatial arrangement of TLVs in both two and three dimensions, we have developed a method for site-selective and sequence-specific sorting of DNA-tagged vesicles to surfaces modified with complementary DNA. The combination of this method with nanoplasmonic sensing formats is covered as well as the possibility of using DNA-modified vesicles for the detection of unlabeled DNA targets on the single-molecule level. Finally, a new method for membrane fusion induced by hybridization of vesicle-anchored DNA is demonstrated, including new results on content mixing obtained with vesicle populations encapsulating short, complementary DNA strands. [ABSTRACT FROM AUTHOR]