Nanoscale surface photovoltage of organic semiconductors with two pass Kelvin probe microscopy.

Kelvin probe microscopy implemented with controlled sample illumination is used to study nanoscale surface photovoltage effects. With this objective a two trace method, where each scanning line is measured with and without external illumination, is proposed. This methodology allows a direct comparison of the contact potential images acquired in darkness and under illumination and, therefore, the surface photovoltage is simply inferred. Combined with an appropriate data analysis, the temporal and spatial evolution of reversible and irreversible photo-induced processes can be obtained. The potential and versatility of this technique is applied to MEH-PPV thin films. Photo-physical phenomena such as the mesoscale polymer electronic light-induced response as well as the local nanoscale electro-optical properties are studied.

AFM imaging and analysis of electrostatic double layer forces on single DNA molecules.

Electrical double layer (EDL) forces develop between charged surfaces immersed in an electrolyte solution. Biological material surrounded by its physiological medium constitutes a case where these forces play a major role. Specifically, this work is focused on the study of the EDL force exerted by DNA molecules, a standard reference for the study of single biomolecules of nanometer size. The molecules deposited on plane substrates have been characterized by means of the atomic force microscope operated in the force spectroscopy imaging mode. Force spectroscopy imaging provides images of the topography of the DNA molecules, and of the EDL force spectrum. Due to the size of the molecule being much smaller than that of the tip, both the tip-substrate and tip-molecule interactions need to be considered in the analysis of the experimental results. We solve this problem by linearly superposing the two contributions. EDL force images are presented where DNA molecules are clearly resolved. The lateral resolution of the EDL force is discussed and compared with that of the topography. The method also allows the estimation of the DNA surface charge density, thereby obtaining reasonable values.

WSXM: a software for scanning probe microscopy and a tool for nanotechnology.

In this work we briefly describe the most relevant features of WSXM, a freeware scanning probe microscopy software based on MS-Windows. The article is structured in three different sections: The introduction is a perspective on the importance of software on scanning probe microscopy. The second section is devoted to describe the general structure of the application; in this section the capabilities of WSXM to read third party files are stressed. Finally, a detailed discussion of some relevant procedures of the software is carried out.

Jumping mode AFM imaging of biomolecules in the repulsive electrical double layer.

We present a method to image single biomolecules in aqueous media by atomic force microscope (AFM) without establishing any mechanical contact between the tip and the sample. It works by placing the feedback set point in the repulsive electrical double-layer curve just before the mechanical instability occurs. We use the jumping operation mode, where the set point is controlled at every image point and a stable imaging is achieved for several hours. This is a necessary condition for this method to be operative, otherwise the tip can fall in contact in a short time. The method is applied to image single-avidin protein molecules deposited on cleaved mica. In addition, the dependence of the height of avidin molecules as a function of ion concentration, due to differences in surface charge density of mica and avidin, is tentatively used to deduce relative values of these quantities.

Atomic force microscopy contact, tapping, and jumping modes for imaging biological samples in liquids.

The capabilities of the atomic force microscope for imaging biomolecules under physiological conditions has been systematically investigated. Contact, dynamic, and jumping modes have been applied to four different biological systems: DNA, purple membrane, Alzheimer paired helical filaments, and the bacteriophage phi29. These samples have been selected to cover a wide variety of biological systems in terms of sizes and substrate contact area, which make them very appropriate for the type of comparative studies carried out in the present work. Although dynamic mode atomic force microscopy is clearly the best choice for imaging soft samples in air, in liquids there is not a leading technique. In liquids, the most appropriate imaging mode depends on the sample characteristics and preparation methods. Contact or dynamic modes are the best choices for imaging molecular assemblies arranged as crystals such as the purple membrane. In this case, the advantage of image acquisition speed predominates over the disadvantage of high lateral or normal force. For imaging individual macromolecules, which are weakly bonded to the substrate, lateral and normal forces are the relevant factors, and hence the jumping mode, an imaging mode which minimizes lateral and normal forces, is preferable to other imaging modes.

Characterization by atomic force microscopy of Alzheimer paired helical filaments under physiological conditions.

Paired helical filaments (PHF) is an aberrant structure present in the brain of Alzheimer's disease patients which has been correlated with their degree of dementia. In order to determine the structure of PHF, several studies have been performed using atomic force microscopy (AFM). However, those studies have the limitation that they have not been done in solution and the sample could be far from the real physiological conditions. In this work we present an AFM analysis of PHF in liquid environment and we compare that analysis with that performed in dry conditions. PHF imaging in liquid was only possible by using jumping mode AFM as the imaging technique. Jumping mode AFM images of PHF in solution show first, a notable increase in the absolute values of the height of the filament, and second, a smaller ratio between the height measured at the upper and at the lower part of the PHF. Direct comparison of the experimental data with structural models has been performed. From this we conclude that the PHF structure is compatible with two coupled ribbons with an overall height of 20 nm and a width of 10 nm.

DNA height in scanning force microscopy.

The measured height of DNA molecules adsorbed on a mica substrate by scanning probe microscopy is always less than the theoretical diameter. In this paper we show that, when imaged in ambient conditions, the molecules are usually immersed in the salt layer used to adsorb them to the substrate. This layer distorts the measurement of DNA height and is the main source of error but not the only one. We have performed different experiments to study this problem using two scanning force techniques: non-contact tapping mode in air and jumping mode in aqueous solution, where the dehydration phenomena is minimized. Height measurements of DNA in air using tapping mode reveal a height of 0.7+/-0.2nm. This value increases up to 1.5+/-0.2nm when the salt layer, in which the molecules are embedded, is removed. Jumping experiments in water give a value of 1.4+/-0.3nm when the maximum applied force is 300pN and 1.8+/-0.2nm at very low forces, which confirms the removal of the salt layer. Still, in all our experiments, the measured height of the DNA is less than the theoretical value. Our results show that although the salt layer present is important, some sample deformation due to either the loading force of the tip or the interaction with the substrate is also present.

Interaction forces and conduction properties between multi wall carbon nanotube tips and Au(111).

We have studied the interaction forces and electrical conduction properties arising between multiwall carbon nanotube tips and the Au(111) surface in air, by means of amplitude modulation scanning force microscopy, also called intermittent contact. We have centered our work on tips with metallic electronic structure and for the specific parameters used we have found a preliminary interaction range where there is no contact between tip and surface. Stable imaging in this non-contact range is possible with multiwall carbon nanotube tips. These tips have also been used to obtain simultaneous topographic and current maps of the surface. They show excellent properties as tips due to their high aspect ratio and durability, as a result of their elastic and non-reactive properties. Correspondingly, multiwall carbon nanotube tips allow high resolution local analysis of electrical conductivity on a nanometer scale.

Contactless experiments on individual DNA molecules show no evidence for molecular wire behavior.

A fundamental requirement for a molecule to be considered a molecular wire (MW) is the ability to transport electrical charge with a reasonably low resistance. We have carried out two experiments that measure first, the charge transfer from an electrode to the molecule, and second, the dielectric response of the MW. The latter experiment requires no contacts to either end of the molecule. From our experiments we conclude that adsorbed individual DNA molecules have a resistivity similar to mica, glass, and silicon oxide substrates. Therefore adsorbed DNA is not a conductor, and it should not be considered as a viable candidate for MW applications. Parallel studies on other nanowires, including single-walled carbon nanotubes, showed conductivity as expected.

Nonlinear resistance versus length in single-walled carbon nanotubes.

In this work fundamental properties of the electrical transport of single-walled carbon nanotubes as a function of their length are investigated. For this purpose, we have developed a new technique that allows us to characterize electronic transport properties of single-walled carbon nanotubes by probing them at different spots. This technique uses scanning force microscopy to make mechanical and electrical nanocontacts at any selected spot of a given image. We have applied this technique to molecules with high intrinsic resistance. The results show a nonlinear resistance vs distance behavior as the nanotube is probed along its length. This is an indication of elastic electronic transport in one-dimensional systems.

Macroscopic water deposits on polycrystalline gold measured by scanning force microscopy.

Data of water adsorption on polycrystalline gold show the formation of a multilayer film of several nanometers with the increase of relative humidity. We have measured this adsorption process by scanning force microscopy in both dynamic and jumping modes. We find interesting differences in the adsorption of water on the terraces and at grain boundaries. Measurements of adhesion force are also reported.

Imaging and mapping protein-binding sites on DNA regulatory regions with atomic force microscopy.

Regulation of gene expression is fundamental in biological systems. A systematic search for protein binding sites in gene promoters has been done in recent years. Biochemical techniques are easy and reliable when analysing protein interactions with short pieces of DNA, but are difficult and tedious when long pieces of DNA have to be analysed. Here we propose AFM as a reliable and easy technique for identifying protein interaction sites in long DNA molecules like gene promoters. We support this idea using a well-known model: the interaction of the Pho4 protein with the PHO5 gene promoter. We have also applied the technique to demonstrate that Mig1 protein binds to two motifs in the promoter of HXK2 gene. Our results allow us to define Mig1p as a new factor probably contributing to the carbon source-dependent transcription regulation of HXK2 gene.

Absence of dc-conductivity in lambda-DNA.

The electrical conductivity of biomaterials on a molecular scale is of fundamental interest in the life sciences. We perform first principles electronic structure calculations, which clearly indicate that lambda-DNA chains should present large resistance values. We also present two direct procedures to measure electrical currents through DNA molecules adsorbed on mica. The lower limit for the resistivity is 10(6) Omega . cm, in agreement with our calculations. We also show that low energy electron bombardment induces a rapid contamination and dramatically affects the measured conductivity, thus providing an explanation to recent reports of high DNA conductivity.

Analysis by atomic force microscopy of Med8 binding to cis-acting regulatory elements of the SUC2 and HXK2 genes of saccharomyces cerevisiae.

Med8 protein is a regulator that specifically binds to upstream activating sequences (UASs) of SUC2 promoter, to downstream repressing sequences (DRSs) of the HXK2 gene and to the carboxy-terminal domain of the RNA polymerase II. Atomic force microscopy has allowed for direct visualization of Med8 interactions with a 305 bp fragment of SUC2 promoter and with a 676 bp fragment of HXK2 gene, containing respectively the UASs and DRSs regulatory regions. This approach has provided complementary information about the position and the structure of the DNA-protein complexes. Med8 binding to DNA results in total covering of one of the two existing 7 bp motives (consensus, (A/C)(A/G)GAAAT) in the studied DNA fragments. No preference for binding either of the two UASs of SUC2 promoter as well as for the two DRSs of HXK2 gene has been found. We also discuss whether this protein works as dimer or as a monomer.

Properties of metallic nanowires: from conductance quantization to localization.

Material structures of reduced dimensions exhibit electrical and mechanical properties different from those in the bulk. Measurements of room-temperature electronic transport in pulled metallic nanowires are presented, demonstrating that the conductance characteristics depend on the length, lateral dimensions, state and degree of disorder, and elongation mechanism of the wire. Conductance during the elongation of short wires (length l approximately 50 angstroms) exhibits periodic quantization steps with characteristic dips, correlating with the order-disorder states of layers of atoms in the wire predicted by molecular dynamics simulations. The resistance R of wires as long as l approximately 400 angstroms exhibits localization characteristics with In R(l) approximately l(2).

Characterization of surface roughness in titanium dental implants measured with scanning tunnelling microscopy at atmospheric pressure.

Characterization of the surface topography of implant materials is important for understanding tissue response. We have measured, for the first time, the topography of titanium surfaces used in osseointegrated dental implants. Scanning tunnelling microscopy (STM) which provides 3D real space images was used. In addition to clinical samples, electropolished and anodically oxidized surfaces were also measured. Clinical samples are rather inhomogeneous in character showing grooves and steps with a maximum depth of 0.11 micron. Micropores with an average diameter of about 30 nm are also present. Electropolished samples are rather homogeneous and very smooth, showing steps of 1 to 5 nm in height. The measurements were performed under atmospheric conditions at a resolution in the subnanometer range.

Determination of surface topography of biological specimens at high resolution by scanning tunnelling microscopy.

Although techniques are available for the determination of the three-dimensional structure of biological specimens, for example scanning electron microscopy, they all have some serious drawback, such as low resolution, the requirement for crystals or for the sample to be analysed in a high vacuum. In an attempt to develop a technique for high-resolution three-dimensional structure analysis of non-crystalline biological material, we have tested the applicability of scanning tunnelling microscopy (STM), a method that has been used successfully in the analysis of metal and semiconductor surface structures. We report here that scanning tunnelling electron microscopy can be used to determine the surface topography of biological specimens at atmospheric pressure and room temperature, giving a vertical resolution of the order of 1 A. Our results show that quantum mechanical tunnelling of electrons through biological material is possible provided that the specimen is deposited on a conducting surface.