Georg Papastavrou

Close Approximation of Two Platelet Factor 4 Tetramers by Charge Neutralization Forms the Antigens Recognized by HIT Antibodies

Objective—Heparin-induced thrombocytopenia (HIT) is a prothrombotic drug reaction caused by antibodies that recognize positively charged platelet factor 4 (PF4), bound to the polyanion, heparin. The resulting immune complexes activate platelets. Unfractionated heparin (UFH) causes HIT more frequently than low-molecular-weight heparin (LMWH), whereas the smallest heparin-like molecule (the pentasaccharide, fondaparinux), induces anti-PF4/heparin antibodies as frequently as LMWH, but without exhibiting cross-reactivity with these antibodies. To better understand these findings, we analyzed the molecular structure of the complexes formed between PF4 and UFH, LMWH, or fondaparinux. Methods and Results—By atomic force microscopy and photon correlation spectroscopy, we show that with any of the 3 polyanions, but in the order, UFH>LMWH≫fondaparinux—PF4 forms clusters in which PF4 tetramers become closely apposed, and to which anti-PF4/heparin antibodies bind. By immunoassay, HIT antibodies bind strongly to PF4/H/PF4 complexes, but only weakly to single PF4/heparin molecules. Conclusion—HIT antigens are formed when charge neutralization by polyanion allows positively charged PF4 tetramers to undergo close approximation. Whereas such a model could explain why all 3 polyanions form antibodies with similar specificities, the striking differences in the relative size and amount of complexes formed likely correspond to the observed differences in immunogenicity (UFH>LMWH≈fondaparinux) and clinically relevant cross-reactivity (UFH>LMWH≫fondaparinux).

Investigating forces between charged particles in the presence of oppositely charged polyelectrolytes with the multi-particle colloidal probe technique.

Direct force measurements are used to obtain a comprehensive picture of interaction forces acting between charged colloidal particles in the presence of oppositely charged polyelectrolytes. These measurements are achieved by the multi-particle colloidal probe technique based on the atomic force microscope (AFM). This novel extension of the classical colloidal probe technique offers three main advantages. First, the technique works in a colloidal suspension with a huge internal surface area of several square meters, which simplifies the precise dosing of the small amounts of the polyelectrolytes needed and makes this approach less sensitive to impurities. Second, the particles are attached in-situ within the fluid cell, which avoids the formation of nanobubbles on the latex particles used. Third, forces between two similar particles from the same batch are being measured, which allows an unambiguous determination of the surface potential due to the symmetry of the system. Based on such direct force measurements involving positively and negatively charged latex particles and different polyelectrolytes, we find the following forces to be relevant. Repulsive electrostatic double-layer forces and attractive van der Waals forces as described by the theory of Derjaguin, Landau, Verwey, and Overbeek (DLVO) are both important in these systems, whereby the electrostatic forces dominate away from the isoelectric point (IEP), while at this point they vanish. Additional non-DLVO attractive forces are operational, and they have been identified to originate from the electrostatic interactions between the patch-charge heterogeneities of the adsorbed polyelectrolyte films. Highly charged polyelectrolytes induce strong patch-charge attractions, which become especially important at low ionic strengths and high molecular mass. More weakly charged polyelectrolytes seem to form more homogeneous films, whereby patch-charge attractions may become negligible. Individual bridging events could be only rarely identified from the retraction part of the force profiles, and therefore we conclude that bridging forces are unimportant in these systems.

Attractive and Repulsive Electrostatic Forces between Positively Charged Latex Particles in the Presence of Anionic Linear Polyelectrolytes

The interaction forces between individual positively charged amidine functionalized latex particles with adsorbed negatively charged sodium poly(styrene sulfonate) were studied with the colloidal probe technique based on atomic force microscopy (AFM). When the polymer dose is progressively increased, the strength of the repulsive force between the particles decreases as the charge neutralization point is approached, then increases again due to overcharging, and finally reaches a plateau. Surface potentials obtained from fits of the force profiles to Poisson-Boltzmann theory agree well with potentials measured with electrophoresis. Close to the charge neutralization point, attractive forces exceeding van der Waals interactions are found. These attractive forces increase in strength with increasing molecular mass of the polymer and decreasing ionic strength. These attractive interactions are of electrostatic origin and result from lateral patch-charge heterogeneities within the adsorbed polyelectrolyte layer. The measured forces are shown to be in semiquantitative agreement with model calculations based on charge distributions with square lattice symmetry.

Attractive Electrostatic Forces between Identical Colloidal Particles Induced by Adsorbed Polyelectrolytes

Polyelectrolytes adsorb strongly at oppositely charged surfaces, thereby dramatically influencing the corresponding interaction forces. In this letter, we report on direct force measurements with the atomic force microscope (AFM) between two individual particles in an aqueous colloidal suspension in the presence of polyelectrolytes near the isoelectric point. From systematic variations of the molecular mass, the ionic strength, and analysis of adhesion events, we conclude that the observed attractive forces are mainly due to electrostatic patch-charge interactions. The same type of attractive forces is equally influencing interactions between proteins as well as hydrophobic or mineral surfaces.

Charge regulation effects on electrostatic patch-charge attraction induced by adsorbed dendrimers

A multi-particle colloidal probe technique based on the atomic force microscope (AFM) was used to measure the interaction forces between individual charged latex particles with adsorbed cationic poly(amido amine) (PAMAM) dendrimers. The forces near the isoelectric point (IEP) were found to be attractive and stronger than van der Waals interactions. These additional attractions can be rationalized semi-quantitatively with a patch-charge model, which is derived from Debye-Hückel theory. However, the model predicts a larger decay constant for the attractive interaction than observed experimentally. The deviation is probably due to the disordered liquid-like structure of the experimentally investigated system and the finite size of the interacting regions. The amplitude of the attractive interaction is estimated correctly by the patch-charge model including its dependence on the ionic strength and dendrimer generation. The experimental force curves are situated between predictions for constant charge and constant potential boundary conditions. We conclude that the observed additional attractive forces are of electrostatic origin, and that charge regulation effects may play an important role.

A Direct Biocombinatorial Strategy toward Next Generation, Mussel-Glue Inspired Saltwater Adhesives

Biological materials exhibit remarkable, purpose-adapted properties that provide a source of inspiration for designing new materials to meet the requirements of future applications. For instance, marine mussels are able to attach to a broad spectrum of hard surfaces under hostile conditions. Controlling wet-adhesion of synthetic macromolecules by analogue processes promises to strongly impact materials sciences by offering advanced coatings, adhesives, and glues. The de novo design of macromolecules to mimic complex aspects of mussel adhesion still constitutes a challenge. Phage display allows material scientists to design specifically interacting molecules with tailored affinity to material surfaces. Here, we report on the integration of enzymatic processing steps into phage display biopanning to expand the biocombinatorial procedure and enable the direct selection of enzymatically activable peptide adhesion domains. Adsorption isotherms and single molecule force spectroscopy show that those de novo peptides mimic complex aspects of bioadhesion, such as enzymatic activation (by tyrosinase), the switchability from weak to strong binders, and adsorption under hostile saltwater conditions. Furthermore, peptide-poly(ethylene oxide) conjugates are synthesized to generate protective coatings, which possess anti-fouling properties and suppress irreversible interactions with blood-plasma protein cocktails. The extended phage display procedure provides a generic way to non-natural peptide adhesion domains, which not only mimic nature but also improve biological sequence sections extractable from mussel-glue proteins. The de novo peptides manage to combine several tasks in a minimal 12-mer sequence and thus pave the way to overcome major challenges of technical wet glues.

Interaction and structure of surfaces coated by poly(vinyl amines) of different line charge densities

Interactions between preadsorbed films of poly(vinyl amine) (PVA) of two different line charge densities on silica substrates were studied with the colloidal probe technique based on the atomic force microscope (AFM). The preadsorbed films were prepared by adsorption of PVA from a pH 4 solution without any added salt. The highly charged PVA adsorbs in a flat configuration and in laterally heterogeneous layers, while the more weakly charged PVA analog adsorbs in thicker and more homogeneous films. As revealed by reflectivity measurements, such preadsorbed PVA films are stable in polyelectrolyte-free solutions. However, force measurements with the colloidal probe reveal that their interactions depend strongly on the ionic strength. Upon approach, interactions are dominated by electrostatic diffuse layer overlap forces. Both PVA films have very similar diffuse layer charge densities of about 1.5 mC/m2. Since these values are substantially lower than what would be expected from the total charge of the adsorbed polyelectrolytes measured by reflectivity, we infer that coadsorption of anions represents the principal mechanism in charge neutralization. Upon retraction, the adhesion between the films is dominated by bridging forces due to single polymer chains. Such bridging adhesion becomes progressively important with increasing ionic strength, whereby their range and frequency increase. The work of adhesion due to bridging is about 0.3 mN/m. At low ionic strengths, the films behave differently. While the highly charged PVA shows unspecific adhesion at small distances, the more weakly charged PVA analog shows few adhesion events occurring at long distances.

Long-ranged attractive forces induced by adsorbed dendrimers: direct force measurements and computer simulations.

Interaction forces between charged interfaces in the presence of oppositely charged dendrimers are studied by experiment and simulation. The experiments involve direct force measurements with an atomic force microscope (AFM) between two negatively charged colloidal particles in the presence of adsorbed, positively charged globular dendrimers. The simulations are carried out by treating the macroions explicitly, while the small salt ions are treated implicitly through the Debye-Huckel approximation. The system undergoes overcharging, and at the isoelectric point long-ranged attractive electrostatic forces are present. The range of the attraction is on the order of half the Debye length at high salt concentration, but it becomes smaller at low salt concentration. Away from the isoelectric point, repulsive electrostatic forces are observed due to diffuse layer overlap. A semiquantitative agreement between experiment and simulation is obtained, despite the fact that the simple theoretical model does not involve any adjustable parameters. This study provides for the first time detailed comparison between experimental and simulation data of interaction forces between colloidal particles in the presence of multivalent macroions and monovalent salt ions.

Space-Resolved In-Plane Moduli of Graphene Oxide and Chemically Derived Graphene Applying a Simple Wrinkling Procedure

Almost 50 years after its fi rst description in 1962 by H.P. Boehm, [ 1 ] the unique properties of graphene, a single carbon layer of the graphitic structure, [ 2 ] were unraveled by A. K. Geim and K. S. Novoselov [ 3 ] who were awarded the Nobel prize for physics in 2010. Besides the novel electronic and thermal properties, the exceptionally high mechanical strength triggered various applications as reinforcing fi llers in nanocomposites, [ 4,5 ] and for fabrication of carbon-based paper-like [ 6 ] and fi brous materials. [ 7 ] The only established up-scalable procedure for graphene production is via reduction of graphene oxide (GO). [ 8–9 ] The process implies several steps: oxidation of bulk graphite to graphite oxide, delamination of graphite oxide into GO by osmotic swelling, and fi nally reduction of GO to chemically derived graphene. Conditions for the oxidation of graphite are rather harsh and, due to the heterogeneous character of the oxidation, gradients in the local concentration of reactants are inherent to the process. Accordingly, the degree of oxidation achieved is heterogeneous within the material and, moreover, critically depends on the kinetics of the reaction. The type and number of functional groups introduced/removed by the redoxchemistry vary spatially. Consequently, properties of GO and graphene derived thereof vary from sample to sample and even within domains of a single nanoplatelet. The functional groups act as local defects and, for instance, electrical conductivity is critically infl uenced by defect concentration and distribution. [ 10 ]

Interpretation of adhesion force between self-assembled monolayers measured by chemical force microscopy

Abstract We show that the theory of the surface energy of condensed phases, which is based on its separation into London–van der Waals and acid–base components, can be used for the interpretation of adhesion force and work of adhesion measurements with the chemical force microscope. However, it can be done in a straightforward manner only for surfaces and liquids with the same type of solvation. Even in this case, some complicating factors like dissimilarity of the interacting hydro- and fluorocarbon groups has to be considered. At lyophilic surfaces liquid tends to form layered solvation structures at the interface and the presence of these structures may influence interfacial energies. We show that this type of effect can be very significant for hydrophilic surfaces in water. For instance, we found the surface free energy of the OH or COOH terminated monolayers in water is 20÷40 mJ m −2 higher than interfacial energy in hexadecane. Additionally, we predict that if liquid forms a layered structure at the interface, the observed adhesion force depends on the load applied to the CFM cantilever. The experimental evidence of such a behavior is presented.