A novel strategy to construct a flat-lying DNA monolayer on a mica surface

Flat-lying, densely packed DNA monolayers in which DNA chains are well organized have been successfully constructed on a mica surface by dropping a droplet of a DNA solution on a freshly cleaved mica surface and subsequently transferring the mica to ultrapure water for developing. The formation kinetics of such monolayers was studied by tapping mode atomic force microscopy (TMAFM) technique. A series of TMAFM images of DNA films obtained at various developing times show that before the sample was immersed into water for developing the DNA chains always seriously aggregated by contacting, crossing, or overlapping and formed large-scale networks on the mica surface. During developing, the fibers of DNA networks gradually dispersed into many smaller fibers up to single DNA chains. At the same time, the fibers or DNA chains also experienced rearrangement to decrease electrostatic repulsion and interfacial Gibbs free energy. Finally, a flat-lying, densely packed DNA monolayer was formed. A formation mechanism of the DNA monolayers was proposed that consists of aggregation, dispersion, and rearrangement. The effects of both DNA and Mg2+ concentration in the formation solution on DNA monolayer formation were also investigated in detail.     

Effects of DNA topology, temperature and solvent viscosity on DNA retardation in slalom chromatography

Slalom chromatography is a unique size-fractionation method applicable to large DNA molecules [>5 kilobase pairs (kbp)]. The method was first developed by using columns packed with microbeads (diameter, <20 microm) used for high-performance liquid chromatography and by applying a relatively fast flow-rate (>0.3 ml/min). Previous studies suggested that the separation is attributed to a hydrodynamic rather than to an equilibrium phenomenon (J. Hirabayashi and K. Kasai, Anal. Biochem. 178 (1989) 336; J. Hirabayashi, N. Itoh, K. Noguchi and K. Kasai, Biochemistry, 29 (1990) 9515). In the present report, the results of a systematic study on the effects of DNA topology, temperature, and solvent viscosity on DNA retardation are described. Firstly, the behaviour of circular (super-coiled) and linearized forms of charomid DNAs (20-42 kbp) was studied. Circular-form DNA molecules were found to be fractionated size-dependently similarly to linear forms in a flow-rate dependent manner. However, the extent of retardation of the circular form DNA was apparently less than that of the corresponding linear forms. Circular DNAs showed almost the same retardation (e.g., 42 kbp) as DNA fragments (e.g., 20 kbp) having approximately half of the size of the former. This observation indicates that DNA retardation is basically related to physical length, not to mass. Secondly, to study the effect of temperature with special reference to solvent viscosity, we carried out chromatographic analysis at various temperatures ranging from 6 to 65 degrees C in both the absence and presence of sucrose (10 or 20%, w/v). The results showed that it is the solvent viscosity that determines the extent of retardation. Taken together, all of physicochemical parameters that define hydrodynamic properties, i.e., particle size, flow-rate and solvent viscosity, proved to be critical in slalom chromatography as well as the potential physical length of the DNA, thus supporting the concept that slalom chromatography is based on a hydrodynamic principle.     

Carbenium ions,onium ions,and covalent species in the cationic polymerization of alkenes

The importance of exchange reactions between various types of active species present in the cationic polymerization of alkenes is addressed. Reactivities of carbocations, corresponding ion pairs, onium ions, and covalent species are discussed for model systems and in real polymerization. The influence of various parameters on the equilibria between carbenium ions, onium ions, and covalent species is described. Rate constants of the isomerization of active sites are estimated and correlated with polydispersities. General ways to improve control of cationic polymerization of alkenes are discussed.     

Effect of bicarbonate ions on the crystallization of calcite on self-assembled monolayers

We use molecular dynamics simulations to investigate the nucleation of calcite crystals on self-assembled monolayers. We show how the presence of bicarbonate ions adsorbed on the monolayer surface can both aid nucleation and control the orientation of the growth of the crystal. Using a simple model of the nucleation process and calculated interfacial energies, we calculate the enhancement (with respect to the homogeneous nucleation rate) of the nucleation of calcite on the (012) and (0001) faces. The calculations show clearly that the (012) face is favored over the (0001) face and that the nucleation rate is enhanced for self-assembled monolayers made from molecules containing an even number of carbon atoms in the alkyl chain over those containing an odd number.     

Effects of liquid bridge between colloidal spheres and evaporation temperature on fabrication of colloidal multilayers

For the application of colloidal crystal films as "photonic band gap" materials, their domain size and thickness are significant. The substrate withdrawing speed, the colloidal suspension volume fraction, and the colloidal suspension temperature have been studied for the domain size and thickness controls of colloidal crystals in this study. Stable dispersions of monodispersed polystyrene spheres with a diameter of 245 nm were synthesized according to a general emulsion polymerization for colloidal crystal films. By experimental results and the theoretical relationship between the number of layers and other parameters, we could know that the water bridge between colloidal spheres (which is formed by capillary force) influences the number of colloidal crystal layers significantly.     

DNA-directed protein immobilization on mixed self-assembled monolayers via a streptavidin bridge

The simultaneous detection of multiple analytes is an important consideration for the advancement of biosensor technology. Currently, few sensor systems possess the capability to accurately and precisely detect multiple antigens. This work presents a simple approach for the functionalization of sensor surfaces suitable for multichannel detection. This approach utilizes self-assembled monolayer (SAM) chemistry to create a nonfouling, functional sensor platform based on biotinylated single-stranded DNA immobilized via a streptavidin bridge to a mixed SAM of biotinylated alkanethiol and oligo(ethylene glycol). Nonspecific binding is minimized with the nonfouling background of the sensor surface. A usable protein chip is generated by applying protein-DNA conjugates which are directed to specific sites on the sensor chip surface by utilizing the specificity of DNA hybridization. The described platform is demonstrated in a custom-built surface plasmon resonance biosensor. The detection capabilities of a sensor using this protein array have been characterized using human chorionic gonadotropin (hCG). The platform shows a higher sensitivity in detection of hCG than that observed using biotinylated antibodies. Results also show excellent specificity in protein immobilization to the proper locations in the array. The vast number of possible DNA sequences combine with the selectivity of base-pairing makes this platform an excellent candidate for a sensor capable of multichannel protein detection.     

Impact of chain length, temperature, and humidity on the growth of long alkyltrichlorosilane self-assembled monolayers

In this work, we have studied the growth of self-assembled monolayers (SAMs) on silicon dioxide (SiO(2)) made of various long alkyltrichlorosilane chains (16, 18, 20, 24, and 30 carbon atoms in the alkyl chain), at several values of temperature (11 and 20 °C in most cases) and relative humidity (18 and 45% RH). Using atomic force microscopy analysis, thickness measurements by ellipsometry, and contact angle measurements, we have built a model of growth behaviour of SAMs of those molecules according to the deposition conditions and the chain length. Particularly, this work brings not only a better knowledge of the less studied growth of triacontyltrichlorosilane (C(30)H(61)SiCl(3)) SAMs but also new results on SAMs of tetracosyltrichlorosilane (C(24)H(49)SiCl(3)) that have not already been studied to our knowledge. We have shown that the SAM growth behaviour of triacontyltrichlorosilane at 20 °C and 45% RH is similar to that obtained at 11 °C and 45% RH for shorter molecules of hexadecyltrichlorosilane (C(16)H(33)SiCl(3)), octadecyltrichlorosilane (C(18)H(37)SiCl(3)), eicosyltrichlorosilane (C(20)H(41)SiCl(3)) and tetracosyltrichlorosilane (C(24)H(49)SiCl(3)). We have also observed that the monolayers grow faster at 45% than at 18% RH, and surprisingly slower at 20 °C than at 11 °C. Another important result is that the growth time constant decreases with the number of carbon atoms in the alkyl chain except for C(24)H(49)SiCl(3) at 11 °C and 18% RH, and for C(30)H(61)SiCl(3). To our knowledge, such a chain length dependence of the growth time constant has never been reported. The latter and all the other results are interpreted by adapting a diffusion limited aggregation growth model.     

Preparation and structure of a low-density, flat-lying decanethiol monolayer from the densely packed, upright monolayer on gold

This study investigates the formation of low-density, flat-lying decanethiol chemisorbed on Au prepared by heating the surface covered with a densely packed, upright monolayer to a surface temperature above that of the onset of desorption. We determined conditions for preparing the low-density phase by observing the evolution of the photoemission spectrum as a function of the surface temperature using polarized ultraviolet light and by utilizing scanning tunneling microscopy. The preparation conditions were similar for single- and polycrystalline gold surfaces. Once the low-density decanethiol phase was formed, reflection absorption infrared spectroscopy was employed to determine the orientation of the carbon chain backbone with respect to the Au surface. The nature of the valance electronic structure for flat-lying decanethiol is described.     

Scanning electrochemical microscopy. 51. Studies of self-assembled monolayers of DNA in the absence and presence of metal ions

Scanning electrochemical microscopy was used to examine electron transfer across a self-assembled monolayer of thiol-modified DNA duplexes on a gold electrode. The apparent rate constant for heterogeneous ET from a solution redox probe, Fe(CN)6(3-/4-), to the gold surface through ds-DNA was 4.6 (+/-0.2) x 10(-7) cm/s. With the addition of Zn2+, which resulted in the formation of a metalated DNA (M-DNA) monolayer, the rate constant increased to 5.0 (+/-0.3) x 10(-6) cm/s. Upon treating M-DNA with EDTA, the zinc ions were released from the monolayer and the original rate constant for the DNA duplexes was restored. The enhanced ET rate was also observed at a DNA monolayer treated with Ca2+ or Mg2+, which does not complex by the DNA bases to form M-DNA. The binding of these cations facilitated the monolayer penetration by the probe mediator Fe(CN)6(3-/4-) and accordingly caused an increased redox signal of the mediator at the ds-DNA-modified electrode. Cationic or neutral mediators were not blocked by the ds-DNA monolayer. These results suggest that although the increased electron transport through M-DNA could partially be ascribed to the intrinsic enhancement of electric conductivity of M-DNA, which has been confirmed by photochemical studies, the change in the surface charge of DNA monolayers on the electrode caused by the binding of metal ions to DNA molecules may play a more important role in the enhancement of current with M-DNA.     

Effects of concentration and temperature on SDS monolayers at the air-solution interface studied by infrared external reflection spectroscopy

Infrared external reflection (IER) spectra of sodium dodecyl sulfate (SDS) monolayers at the air-solution interface and infrared transmission spectra of the corresponding aqueous solutions were measured at various SDS concentrations and temperatures. A comparison between the spectra of adsorbed monolayers and bulk solutions revealed that the conformational order of the SDS alkyl-chain at the air-solution interface improved with increasing the SDS concentrations, up until the saturation adsorption, and that the conformational order of the adsorbed SDS monolayer was higher than those of monomers and micelles. In addition, below the Krafft point temperature, the adsorbed SDS was maintained in the liquid crystal state, while SDS in the bulk solution was in the crystalline state. Furthermore, the SDS adsorption density was evaluated based on the IER band intensities of the insoluble monolayer of tridecanoic acid with an identical alkyl chain length to SDS.     

Selective electrochemical recognition of ions in solution and at self-assembled monolayers

Quinone-functionalized calix[4]arenes having carboxylic acid groups or thiol groups were prepared and their spontaneous adsorption on silver and gold surfaces, respectively, was studied. Since the cavity-like structure of calixarenes was immobilized on the noble metal electrodes, they exhibited a selective affinity towards specific hard metal ions in aqueous media. Voltammetric and spectroscopic studies showed the well-ordered deposition of organic receptors and entrapment of metal ions. It also was found that the repeated capture and removal of metal ions reversibly with chelating agents such as ethylenediaminetetraacetic acid (EDTA) was possible. This is the first example, to our knowledge, of voltammetric detection of hard metal ions in aqueous media using a chemically modified electrode with redox-active macrocyclic receptors.     

Reaction of 5-40 eV ions with self-assembled monolayers

The reaction of 5-40 eV O(+) and Ne(+) ions with alkanethiolate and semifluorinated alkanethiolate self-assembled monolayers (SAMs) is studied under ultrahigh vacuum (UHV) conditions. Whereas Ne(+) simply sputters fragments from the surface, O(+) can also abstract surface atoms and break C-C bonds in both the hydrocarbon and fluorocarbon SAM chains. Isotopic labeling experiments reveal that O(+) initially abstracts hydrogen atoms from the outermost two carbon atoms on an alkanethiolate SAM chain. However, the position of the isotopic label quickly becomes scrambled along the chain as the SAM is damaged through continuous ion bombardment. Scanning tunneling microscopy (STM) monitors changes in the SAM conformational structure at various stages during 5 eV ion bombardment. STM images indicate that O(+) reacts less efficiently with dodecanethiolate molecules packed internally within a structural domain than it does with molecules adsorbed at domain boundaries or near defect sites. STM images recorded after Ne(+) bombardment suggest that Ne(+) attacks the SAM exclusively near the domain boundaries. Taken collectively, these experiments advance our understanding of the degradation pathways suffered by polymeric satellite materials in the low-earth orbit (LEO) space environment.     

Effects of metal ions on concentration of DNA in high-conductivity media by capillary electrophoresis

A procedure for the rapid determination of mono-, di- and tributyltin in water samples is described. The analytes are simultaneously ethylated and concentrated on a solid-phase microextraction fibre placed in the headspace over the sample for 2 min. The ethylated species are then separated and selectively quantified in only 90 s using a multicapillary gas chromatography column combined with atomic emission detection. The influence of blank signals and sampling conditions on the sensitivity of the method is described. Detection limits of 1-5 ng/l and relative standard deviations of 6-10% at concentrations of 20 ng/l were obtained.     

Electrochemical genosensor based on three-dimensional DNA polymer brushes monolayers

A new approach to the three dimensional integration of short DNA strands at gold electrode surfaces via the in situ formation of DNA-acrylamide conjugates is presented. Surface initiated atom transfer radical polymerisation was employed to grow acrylamide brushes co-polymerised in the presence of acrylamide modified DNA probes. This strategy was demonstrated for the realisation of biofunctionalised thin polymer films capable of binding its complementary 105-base DNA amplicon. The synthesised brushes were characterised using atomic force microscopy, attenuated total reflectance spectroscopy and electrochemical impedance spectroscopy. Once characterised, the polymer brushes were applied to the quantitative detection of target DNA using an enzyme labelled reporter DNA probe in a sandwich-type format.     

The interaction of double-charged metal ions with monolayers and bilayers of phospholipids

Electrophoretic mobilities of hexadecane/water emulsions containing dimyristoyl-phosphatidylcholine (DMPC) or egg yolk lecithin (EYL) monolayers at the interface and those of liposomes prepared from the same lipids were measured as functions of the concentrations of Ca²⁺, Mn²⁺, Cu²⁺, and Ni²⁺ cations in the aqueous phase. The surface potentials, surface charge densities (σ), and the Langmuir adsorption isotherms for various distances from the charged surface to the slip plane (d) were calculated on the basis of the Gouy-Chapman theory for 1∶2 electrolytes and the values of ζ-potentials. The binding constants (K) and parametersd were determined under the assumption that the maximum σ values correspond to one ion per phospholipid molecule at the interface. In the case of DMPC, the ion binding constants (L mol⁻¹) at 25°C are 230 and 87 for Ca²⁺, 31.5 and 21 for Mn²⁺, 11 and 6 for Cu²⁺, and 7.5 and 5.3 for Ni²⁺ in liposomes and emulsions, respectively. The affinities of Cu²⁺ and Ni²⁺ ions for EYL monolayers and bilayers are lower than those for DMPC mono- and bilayers. Thed parameters for all ions are smaller than the radii of the hydrated ions. In the case of Ca²⁺, Cu²⁺, and Ni²⁺, thed values for mono- and bilayers are different. The differences in K values between monolayers and bilayers as well as those between DMPC and EYL mono- and bilayers can be attributed to the differences in the local environment and orientation of the interfacial phosphate groups in these systems. Translated fromIzvestiya Akademii Nauk. Seriya Khimicheskaya, No. 12, pp. 2490–2495, December, 1998.     

Nanomechanics of the formation of DNA self-assembled monolayers and hybridization on microcantilevers

Biomolecular interactions over the surface of a microcantilever can produce its bending motion via changes of the surface stress, which is referred to nanomechanical response. Here, we have studied the interaction forces responsible for the bending motion during the formation of a self-assembled monolayer of thiolated 27-mer single-stranded DNA on the gold-coated side of a microcantilever and during the subsequent hybridization with the complementary nucleic acid. The immobilization of the single-stranded DNA probe gives a mean surface stress of 25 mN/m and a mean bending of 23 nm for microcantilevers with a length and thickness of about 200 microm and 0.8 microm, respectively. The hybridization with the complementary sequence could not be inferred from the nanomechanical response. The nanomechanical response was compared with data from well-established techniques such as surface plasmon resonance and radiolabeling, to determine the surface coverage and study the intermolecular forces between neighboring DNA molecules anchored to the microcantilever surface. From both techniques, an immobilization surface density of 3 x 10(12) molecules/cm(2) and a hybridization efficiency of 40% were determined. More importantly, label-free hybridization was clearly detected in the same conditions with a conventional sensor based on surface plasmon resonance. The results imply that the nanomechanical signal during the immobilization process arises mainly from the covalent attachment to the gold surface, and the interchain interactions between neighboring DNA molecules are weak, producing an undetectable surface stress. We conclude that detection of nucleic acid hybridization with nanomechanical sensors requires reference cantilevers to remove nonspecific signals, more sensitive microcantilever geometries, and immobilization chemistries specially addressed to enhance the surface stress variations.