Discrimination of Adsorbed Double-Stranded and Single-Stranded DNA Molecules on Surfaces by Fluorescence Emission Spectroscopy Using Acridine Orange Dye.

A sensitive and straightforward method for discriminating between surface-adsorbed double-stranded DNA (dsDNA) and single-stranded DNA (ssDNA), based on analysis of the fluorescence emission spectra of DNAs dyed with the metachromatic dye acridine orange, has been developed. Since the degree of discrimination between dsDNA and ssDNA is dependent on dye-base ratio (as has been shown in early studies of DNAs in solution), a specific, reproducible protocol for obtaining good ss-ds discrimination was needed. We studied the emission spectra for DNAs dyed in-situ on two different surfaces, polymethylmethacrylate and poly-l-lysine, using acridine orange solutions of varying concentrations in either 2-(N-morpholino)ethanesulfonic acid (MES) or Tris-Borate EDTA (TBE) buffers. The method should prove useful in characterizing the efficacy of denaturing techniques applied to surface-adsorbed DNAs in preparation for hybridization, replication and transcription experiments on stretched and aligned DNAs.

Reduced viscosity of the free surface in entangled polymer melt films.

By embedding "dilute" gold nanoparticles in single polystyrene thin films as "markers", we probe the local viscosity of the free surface at temperatures far above the glass transition temperature (T(g)). The technique used was x-ray photon correlation spectroscopy with resonance-enhanced x-ray scattering. The results clearly showed the surface viscosity is about 30% lower than the rest of the film. We found that this reduction is strongly associated with chain entanglements at the free surface rather than the reduction in T(g).

The use of graft copolymers to bind immiscible blends.

Computer simulations and experimental studies were combined to design copolymers that enhance the strength of polymer composites. These copolymers contain side chains that associate across the boundary between phase-separated regions to form a "molecular velcro" that effectively binds the regions together. This behavior significantly improves the structural integrity and mechanical properties of the material. Because the side chains can be fabricated from a large class of compounds, the technique greatly increases the variety of copolymers that can be used in forming high-strength polymer blends.

Supercritical fluid processing of polymer thin films: an X-ray study of molecular-level porosity.

This paper reviews our recent experimental results that address the effects of solvent density inhomogeneities in supercritical carbon dioxide (scCO(2)) on polymer thin film processing. The key phenomenon is excess sorption of CO(2) molecules into polymer thin films even when the bulk miscibility with CO(2) is very poor. We have found that the amount of the excess sorption is attributed to the large density fluctuations in scCO(2) near the critical point. Further, taking advantage of the vitrification process of polymer chains through quick evaporation of CO(2), we can preserve the "expanded" structures as they are. The resultant films have large degree of molecular-level porosity that is very useful in producing coatings with low dielectric constants, enhanced adhesion, and metallization properties. These characteristics can be achieved in an environmentally "green" manner, without organic solvents, and are not specific to any class of polymers.

Shear modulation force microscopy study of near surface glass transition temperatures

We report results of glass transition (T(g)) measurements for polymer thin films using atomic force microscopy (AFM). The AFM mode, shear modulation force microscopy (SMFM), involves measuring the temperature-dependent shear force on a tip modulated parallel to the sample surface. Using this method we have measured the surface T(g) of thin (17-500 nm) polymer films and found that T(g) is independent of film thickness (t>17 nm), strength of substrate interactions, or even presence of substrate.

Self-assembled monolayers of rigid thiols.

The preparation, structure, properties and applications of self-assembled monolayers (SAMs) of rigid 4-mercapto-biphenyls are briefly reviewed. The rigid character of the biphenyl moiety results in a molecular dipole moment that affects both the adsorption kinetics on gold surfaces, as well as the equilibrium structure of mixed SAMs. Due to repulsive intermolecular interaction, the Langmuir isotherm model does not fit the adsorption kinetics of these biphenyl thiols, and a new Ising model was developed to fit the kinetics data. The equilibrium structures of SAMs and mixed SAMs depend on the polarity of the solution from which they were assembled. Infrared spectroscopy suggests that biphenyl moieties in SAMs on gold have small tilt angles with respect to the surfaces normal. Wetting studies shows that surfaces of these SAMs are stable for months, thus providing stable model surfaces that can be engineered at the molecular level. Such molecular engineering is important for nucleation and growth studies. The morphology of glycine crystals grown on SAM surfaces depends on the structure of the nucleating glycine layer, which, in turn, depends on the H-bonding of these molecules with the SAM surface. Finally, the adhesion of PDMS cross-linked networks to SAM surfaces depends on the concentration of interfacial H-bonding. This non-linear relationship suggests that the polymeric nature of the elastomer results in a collective H-bonding effect.

Evaluation of fibrinogen self-assembly: role of its αC region.

BACKGROUND: Exposure of cryptic, functional sites on fibrinogen upon its adsorption to hydrophobic surfaces of biomaterials has been linked to an inflammatory response and fibrosis. Such adsorption also induces ordered fibrinogen aggregation which is poorly understood. OBJECTIVE: To investigate hydrophobic surface-induced fibrinogen aggregation. METHODS: Contact and lateral force scanning probe microscopy, yielding topography, image dimensions and fiber elastic modulus measurements were used along with transmission and scanning electron microscopy. Fibrinogen aggregation was induced under non-enzymatic conditions by adsorption on a trioctyl-surface monolayer (trioctylmethylamine) grafted onto silica clay plates. RESULTS: A more than one molecule thick coating was generated by adsorption on the plate from 100 to 200 µg mL⁻¹ fibrinogen solutions, and three-dimensional networks formed from 4 mg mL⁻¹ fibrinogen incubated with uncoated or fibrinogen-coated plates. Fibrils appeared laterally assembled into branching and overlapping fibers whose heights from the surface ranged from approximately 3 to 740 nm. The elastic modulus of fibrinogen fibers was 1.55 MPa. No fibrils formed when fibrinogen lacking αC-domains was used as a coating or was incubated with intact fibrinogen-coated plates, or when the latter plates were sequentially incubated with anti-Aα529-539 mAb and intact fibrinogen. When an anti-Aα241-476 mAb was used instead, fine, long fibers formed. Similarly, sequential incubations of fibrinogen-coated plates with recombinant αC-domain (Aα392-610 fragment) or αC-connector (Aα221-372 fragment) and fibrinogen resulted in distinctly fine fiber networks. CONCLUSIONS: Adsorption-induced fibrinogen self-assembly is initiated by a more than one molecule-thick surface layer and eventuates in three-dimensional networks whose formation requires fibrinogen with intact αC-domains.

Multi-minimum adiabatic potential in the single crystal normal spinel ZnAl(2)O(4), doped by Cu(2+) ions.

Spectroscopic investigations of a ZnAl(2)O(4) spinel doped with bivalent copper ions of 0.05% concentration have been carried out in the temperature range 4.2-290 K using a 3 cm(-1) range electron paramagnetic resonance (EPR) spectrometer having an operational frequency f = (9.241 ± 0.001) GHz. The spectrum can be represented as a superposition of two components: a low-temperature (LT) and a high-temperature (HT) one. Redistribution of integrated intensity between HT and LT components of the spectra occurs with temperature change that is typical of systems with multi-minimum adiabatic potential. Spectra observed are explained within the modified theory of crystalline field (MTCF). The electron levels of a Cu(2+) ion placed in an octahedral coordination center with trigonal distortion [CuO(6)](10-) have been calculated. The influence of possible types of oxygen octahedron distortions and possible displacement of copper ions from the symmetry center on the electron spectrum, as well as the shape of the adiabatic potential, has been analyzed. It is shown that in the low-temperature phase the multiple minima of the adiabatic potential occur due to tetragonal distortions while the depth of a minimum is determined by the degree of trigonal octahedron distortions. Tetragonal distortion values and multi-minimum potential barrier heights have been determined.

Evidence for viscoelastic effects in surface capillary waves of molten polymer films.

The surface dynamics of supported ultrathin polystyrene films with thickness comparable to the radius of gyration were investigated by surface sensitive x-ray photon correlation spectroscopy. We show for the first time that the conventional model of capillary waves on a viscous liquid has to be modified to include the effects of a shear modulus in order to explain both static and dynamic scattering data from ultrathin molten polymer films.

Templated biomineralization on self-assembled protein fibers.

Biological mineralization of tissues in living organisms relies on proteins that preferentially nucleate minerals and control their growth. This process is often referred to as "templating," but this term has become generic, denoting various proposed mineral-organic interactions including both chemical and structural affinities. Here, we present an approach using self-assembled networks of elastin and fibronectin fibers, similar to the extracellular matrix. When induced onto negatively charged sulfonated polystyrene surfaces, these proteins form fiber networks of approximately 10-mum spacing, leaving open regions of disorganized protein between them. We introduce an atomic force microscopy-based technique to measure the elastic modulus of both structured and disorganized protein before and during calcium carbonate mineralization. Mineral-induced thickening and stiffening of the protein fibers during early stages of mineralization is clearly demonstrated, well before discrete mineral crystals are large enough to image by atomic force microscopy. Calcium carbonate stiffens the protein fibers selectively without affecting the regions between them, emphasizing interactions between the mineral and the organized protein fibers. Late-stage observations by optical microscopy and secondary ion mass spectroscopy reveal that Ca is concentrated along the protein fibers and that crystals form preferentially on the fiber crossings. We demonstrate that organized versus unstructured proteins can be assembled mere nanometers apart and probed in identical environments, where mineralization is proved to require the structural organization imposed by fibrillogenesis of the extracellular matrix.

Substrate effect on the melting temperature of thin polyethylene films.

Strong dependence of the crystal orientation, morphology, and melting temperature (Tm) on the substrate is observed in the semicrystalline polyethylene thin films. The Tm decreases with the film thickness decrease when the film is thinner than a certain critical thickness, and the magnitude of the depression increases with increasing surface interaction. We attribute the large Tm depression to the decrease in the overall free energy on melting, which is caused by the substrate attraction force to the chains that competes against the interchain force which drives the chains to crystallization.

Deviations from liquidlike behavior in molten polymer films at interfaces.

We have performed x-ray specular and diffuse scattering on liquid polymer films and analyzed the spectra as a function of film thickness and molecular weight. The results show that films whose molecular weight is close to the entanglement length behave as simple liquids except that the shortest wavelength is determined by the radius of gyration (R(g)) rather than the monomer-monomer distance. When the molecular weight is higher than the entanglement length, the strong deviations from liquidlike behavior are observed. We find that the long wavelength cutoff vector, q(l,c), scales with film thickness, d as d(-1.1+/-0.1) rather than the usual d(-2) expected for simple liquids. If we assume that these deviations are due to surface pinning of the polymer chains, then our results are consistent with the formalism developed by Fredrickson et al. to explain the capillary wave spectrum that can propagate in a polymer brush.

Dynamics of ultrathin films in the glassy state.

We report hole growth experiments in free-standing polystyrene (PS) films at temperatures up to 10 degrees C below the bulk glass transition. The data show an unexpected result: the growth rate of nucleated holes increases with increasing molecular weight, up to a limiting value beyond which the rate is approximately constant. Film thicknesses of 45, 80, and 100 nm were studied, using PS molecular weights ranging from 65K to 11.4 Mg/mol. Hole diameters grew linearly with time, and no growing rims were observed to form around the developing holes. Possible explanations in terms of elasticity, yield, and influence of sample preparation and confinement effects are discussed.

Size-controlled synthesis and characterization of thiol-stabilized gold nanoparticles.

Size-controlled synthesis of nanoparticles of less than a few nanometers in size is a challenge due to the spatial resolution limit of most scattering and imaging techniques used for their structural characterization. We present the self-consistent analysis of the extended x-ray absorption fine-structure (EXAFS) spectroscopy data of ligand-stabilized metal nanoclusters. Our method employs the coordination number truncation and the surface-tension models in order to measure the average diameter and analyze the structure of the nanoparticles. EXAFS analysis was performed on the two series of dodecanethiol-stabilized gold nanoparticles prepared by one-phase and two-phase syntheses where the only control parameter was the gold/thiol ratio xi, varied between 6:1 and 1:6. The two-phase synthesis resulted in the smaller particles whose size decreased monotonically and stabilized at 16 A when xi was lowered below 1:1. This behavior is consistent with the theoretically predicted thermodynamic limit obtained previously in the framework of the spherical drop model of Au nanoparticles.

Effect of density fluctuating supercritical carbon dioxide on polymer interfaces.

We investigated an effect of CO2 sorption on the compatibility of immiscible polystyrene (PS) and polybutadiene (PB) bilayers by using in situ neutron reflectivity. By labeling either polymer with deuterium, we found that the excess CO2 molecules were adsorbed to both top PS and bottom PB layers when the bilayers were exposed to CO2 at the narrow T and P regime near the critical point of pure CO2. Furthermore, we clarified that this excess sorption of CO2 molecules increased the interfacial width between the layers up to 100 angstroms even near room temperature, while the interfacial width without CO2 exposure has been reported to be at most 40 A even at the highest temperature (T congruent with 175 degrees C).

DNA electrophoresis on a flat surface.

We report a new approach for performing DNA electrophoresis. Using experimental studies and molecular dynamics simulations, we show that a perfectly flat silicon wafer, without any surface features, can be used to fractionate DNA in free solution. We determine that the ability of a flat surface to separate DNA molecules results from the local friction between the surface and the adsorbed DNA segments. We control this friction by coating the Si surface with silane monolayer films and show that it is possible to systematically change the size range of DNA that can be separated.

Mobility of polymer chains confined at a free surface.

Dynamic secondary ion mass spectrometry was used to investigate the chain mobility of polystyrene (MW ranging from 4.3 to 957 kg/mol) at the free surface. The data show that the diffusion coefficient was reduced relative to the bulk value within a distance, d < or = 4R(g), from the surface and scaled as 1/N(2.5) at fixed d. These results are in excellent agreement with self-consistent field calculations of the surface segmental distribution and provide the first direct confirmation of various theoretical models that predict asymmetric segmental fluctuation which arises from surface induced orientation of polymer chains.

Thermally induced phase transitions and morphological changes in organoclays.

Thermal transitions and morphological changes in Cloisite organoclays were investigated by differential scanning calorimetry (DSC), thermogravimetric analysis (TGA), Fourier transform infrared (FTIR) spectroscopy, and in situ simultaneous small-angle X-ray scattering (SAXS) and wide-angle X-ray diffraction (WAXD) over the temperature range of 30-260 degrees C. On the basis of DSC and FTIR results, the surfactant component in organoclays was found to undergo a melting-like order-disorder transition between 35 and 50 degrees C. The transition temperatures of the DSC peaks (Ttr) in the organoclays varied slightly with the surfactant content; however, they were significantly lower than the melting temperature of the free surfactant (dimethyldihydrotallowammonium chloride; Tm = 70 degrees C). FTIR results indicated that within the vicinity of Ttr, the gauche content increased significantly in the conformation of surfactant molecules, while WAXD results did not show any change in three-dimensional ordering. Multiple scattering peaks were observed in SAXS profiles. In the SAXS data acquired below Ttr, the second scattering peak was found to occur at an angle lower than twice that of the first peak position (i.e., nonequidistant scattering maxima). In the data acquired above Ttr, the second peak was found to shift toward the equidistant position (the most drastic shift was seen in the system with the highest surfactant content). Using a novel SAXS modeling technique, we suggest that the appearance of nonequidistant SAXS maxima could result from a bimodal layer thickness distribution of the organic layers in organoclays. The occurrence of the equidistant scattering profile above Ttr could be explained by the conversion of the bimodal distribution to the unimodal distribution, indicating a redistribution of the surfactant that is nonbounded to the clay surface. At temperatures above 190 degrees C, the scattering maxima gradually broadened and became nonequidistant again but having the second peak shifted toward a scattering angle higher than twice the first peak position. The changes in SAXS patterns above 190 degrees C could be attributed to the collapse of organic layers due to desorption and/or degradation of surfactant component, which was supported by the TGA data.

X-ray scattering from freestanding polymer films with geometrically curved surfaces.

We show that the x-ray surface scattering from a freestanding polymer film exhibits features that cannot be explained by the usual stochastic formalism for surfaces with random height fluctuations. Instead, a geometric description of the film morphology assuming two curved surfaces characterized by a radius of curvature and a lateral cutoff length successfully accounts for the phase difference between the Kiessig fringes of the nominal "specular" and "off-specular" components of the scattering. The formalism allows one to distinguish unambiguously between conformal and anticonformal curvature morphologies at long length scales.