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Understanding the lateral organization of biological membranes plays a key role on the road to fully appreciate the physiological functions of this fundamental barrier between the inside and outside regions of a cell. Ternary lipid bilayers composed of a high and a low melting temperature lipid and cholesterol represent a model system that mimics some of the important thermodynamical features of much more complex lipid mixtures such as those found in mammal membranes. The phase diagram of these ternary mixtures can be studied exploiting fluorescence microscopy in giant unilamellar vesicles, and it is typically expected to give rise, for specific combinations of composition and temperature, to regions of two-phase coexistence and a region with three-phase coexistence, namely, the liquid-ordered, liquid-disordered, and solid phases. Whereas the observation of two-phase coexistence is routinely possible using fluorescence microscopy, the three-phase region is more elusive to study. In this article, we show that particular lipid mixtures containing diphytanoyl-phosphatidylcholine and cholesterol plus different types of sphingomyelin (SM) are prone to produce bilayer regions with more than two levels of fluorescence intensity. We found that these intensity levels occur at low temperature and are linked to the copresence of long and asymmetric chains in SMs and diphytanoyl-phosphatidylcholine in the lipid mixtures. We discuss the possible interpretations for this observation in terms of bilayer phase organization in the presence of sphingolipids. Additionally, we also show that in some cases, liposomes in the three-phase coexistence state exhibit extreme sensitivity to lateral tension. We hypothesize that the appearance of the different phases is related to the asymmetric structure of SMs and to interdigitation effects.

Antimicrobial peptides (AMPs) and lipopeptides (LPs) represent very promising molecules to fight resistant bacterial infections due to their broad-spectrum of activity, their first target, i.e. the bacterial membrane, and the rapid bactericidal action. For both types of molecules, the action mechanism starts from the membrane of the pathogen agents, producing a disorganization of their phase structure or the formation of pores of different size altering their permeability. This mechanism of action is based on physical interactions more than on a lock-and-key recognition event and it is difficult for the pathogens to rapidly develop an effective resistance. Very small differences in the sequence of both AMPs and LPs might lead to very different effects on the target membrane. Therefore, a correct understanding of their mechanism of action is required with the aim of developing new synthetic peptides, analogues of the natural ones, with specific and more powerful bactericidal activity. Atomic force microscopy (AFM), with its high resolution and the associated force spectroscopy resource, provides a valuable technique to investigate the reorganization of lipid bilayers exposed to antimicrobial or lipopeptides. Here, we present AFM results obtained by ours and other groups on the action of AMPs and LPs on supported lipid bilayers (SLBs) of different composition. We also consider data obtained by fluorescence microscopy to compare the AFM data with another technique which can be used on different lipid bilayer model systems such as SLBs and giant unilamellar vesicles. The outcomes here presented highlight the powerful of AFM-based techniques in detecting nanoscale peptide-membrane interactions and strengthen their use as an exceptional complementary tool to in vivo investigations. Indeed, the combination of these approaches can help decipher the mechanisms of action of different antimicrobials and lipopeptides at both the micro and nanoscale levels, and to design new and more efficient antimicrobial compounds.

An innovative platform for the study of the molecular mechanisms at the basis of mechanotransduction has been implemented, developing an experimental approach capable of providing controlled dynamic compression stimuli and retrieving the biomolecular response with single-cell sensitivity. The system provides the ability to perform compression-release cycles on single cells with controlled forces in the nN range and a user-defined repetition rate. Experimental procedures to perform qPCR from a small set of single cells were finely tuned. The experimental platform was tested in the context of bone (cell line hFOB 1.19), a physiological environment highly subjected to mechanical stimuli. Target genes were identified in the literature, based on their involvement in the osteogenesis process or in the bone response to mechanical stimuli. qPCR analysis shows an increase in expression of the chosen targets, and confirms the effectiveness of the presented approach for studying living single cells response to dynamic compression.

Protein misfolding and aggregation are the common mechanisms in a variety of aggregation-dependent diseases. The compromised proteins often assemble into toxic, accumulating amyloid-like structures of various lengths and their toxicity can also be transferred both in vivo and in vitro a prion-like behavior. The characterization of protein interactions, degradation and conformational dynamics in biological systems still represents an analytical challenge in the prion-like protein comprehension. In our work, we investigated the nature of a transferable cytotoxic agent, presumably a misfolded protein, through the coupling of a multi-detector, non-destructive separation platform based on hollow-fiber flow field-flow fractionation with imaging and downstream in vitro tests. After purification with ion exchange chromatography, the transferable cytotoxic agentwas analyzed with Atomic Force Microscopy and statistical analysis, showing that the concentration of protein dimers and low n-oligomer forms was higher in the cytotoxic sample than in the control preparation. To assess whether the presence of these species was the actual toxic and/or self-propagating factor, we employed HF5 fractionation, with UV and Multi-Angle Light Scattering detection, to define proteins molar mass distribution and abundance, and fractionate the sample into size-homogeneous fractions. These fractions were then tested individually in vitro to investigate the direct correlation with cytotoxicity. Only the later-eluted fraction, which contains high-molar mass aggregates, proved to be toxic onto cell cultures. Moreover, it was observed that the selective transfer of toxicity also occurs for one lower-mass fraction, suggesting that two different mechanisms, acute and later induced toxicity, are in place. These results strongly encourage the efficacy of this platform to enable the identification of protein toxicants.

Nucleic acid (NA) extraction is a basic step for genetic analysis, from scientific research to diagnostic and forensic applications. It aims at preparing samples for its application with biomolecular technologies such as isothermal and non-isothermal amplification, hybridization, electrophoresis, Sanger sequencing and next-generation sequencing. Multiple steps are involved in NA collection from raw samples, including cell separation from the rest of the specimen, cell lysis, NA isolation and release. Typically, this process needs molecular biology facilities, specialized instrumentation and labor-intensive operations. Microfluidic devices have been developed to analyze NA samples with high efficacy and sensitivity. In this context, the integration within the chip of the sample preparation phase is crucial to leverage the promise of portable, fast, user-friendly and economic point-of-care solutions. This review presents an overview of existing lab-on-a-chip (LOC) solutions designed to provide automated NA extraction from human raw biological fluids, such as whole blood, excreta (urine and feces), saliva. It mainly focuses on LOC implementation aspects, aiming to describe a detailed panorama of strategies implemented for different human raw sample preparations.

The present tutorial is aimed at introducing the reader to the main aspects of electron transfer in nanobiodevices. Nanobiodevices are faced both from scientific and technological viewpoints and their particular implementation as electron transfer devices provides the opportunity of presenting fundamentals of electron transfer theory. Examples of implementations of stand alone devices, along with those involving reconfigurable set-ups based on an electrochemical scanning tunneling microscope, enable introducing heterogeneous electron transfer and electron transport theories in electrochemical environment. Specific cases of nanobiodevices involving redox metalloproteins are reported and experimental results are interpreted and discussed in view of the most recent theoretical advancements, in order to provide the reader with a comprehensive view of the results and promises in this exciting branch of nanotechnology.

The ability to sense mechanical stimuli and elaborate a response to them is a fundamental process in all organisms, driving crucial mechanisms ranging from cell volume regulation up to organ development or regeneration. Nevertheless, only in few cases the underlying molecular players are known. In particular, mammals possess a large variety of mechanoreceptors, providing highly specialized functions in sensory cells, but also several housekeeping molecular systems are involved in the complex mechanism of mechanotransduction. Recently, a new class of almost ubiquitous membrane channels has been identified in mammalians, namely piezo1 and piezo2, that is thought to play a crucial role in the mechanobiology of mammals. This review focuses on recent findings on these novel channels, and highlights open biophysical questions that largely remain to be addressed.

Amphiphilic molecules supposed to affect membrane protein activity could strongly interact also with the lipid component of the membrane itself. Neurosteroids are amphiphilic molecules that bind to plasma membrane receptors of cells in the central nervous system but their effect on membrane is still under debate. For this reason it is interesting to investigate their effects on pure lipid bilayers as model systems. Using the micropipette aspiration technique (MAT), here we studied the effects of a neurosteroid, allopregnanolone (3?,5?-tetrahydroprogesterone or Allo) and of one of its isoforms, isoallopregnanolone (3?,5?-tetrahydroprogesterone or isoAllo), on the physical properties of pure lipid bilayers composed by DOPC/bSM/chol. Allo is a well-known positive allosteric modulator of GABAA receptor activity while isoAllo acts as a non-competitive functional antagonist of Allo modulation. We found that Allo, when applied at nanomolar concentrations (50-200 nM) to a lipid bilayer model system including cholesterol, induces an increase of the lipid bilayer area and a decrease of the mechanical parameters. Conversely, isoAllo, decreases the lipid bilayer area and, when applied, at the same nanomolar concentrations, it does not affect significantly its mechanical parameters. We characterized the kinetics of Allo uptake by the lipid bilayer and we also discussed its aspects in relation to the slow kinetics of Allo gating effects on GABAA receptors. The overall results presented here show that a correlation exists between the modulation of Allo and isoAllo of GABAA receptor activity and their effects on a lipid bilayer model system containing cholesterol.

Amphiphilic molecules which have a biological effect on specific membrane proteins, could also affect lipid bilayer properties possibly resulting in a modulation of the overall membrane behavior. In light of this consideration, it is important to study the possible effects of amphiphilic molecule of pharmacological interest on model systems which recapitulate some of the main properties of the biological plasma membranes. In this work we studied the effect of a neurosteroid, Allopregnanolone (3?,5?-tetrahydroprogesterone or Allo), on a model bilayer composed by the ternary lipid mixture DOPC/bSM/chol. We chose ternary mixtures which present, at room temperature, a phase coexistence of liquid ordered (Lo) and liquid disordered (Ld) domains and which reside near to a critical point. We found that Allo, which is able to strongly partition in the lipid bilayer, induces a marked increase in the bilayer area and modifies the relative proportion of the two phases favoring the Ld phase. We also found that the neurosteroid shifts the miscibility temperature to higher values in a way similarly to what happens when the cholesterol concentration is decreased. Interestingly, an isoform of Allo, isoAllopregnanolone (3?,5?-tetrahydroprogesterone or isoAllo), known to inhibit the effects of Allo on GABAA receptors, has an opposite effect on the bilayer properties.

We have devised a supramolecular edifice involving His-tagged protein A and antibodies to yield surface immobilized, uniformly oriented, IgG-type, antibody layers with Fab fragments exposed off an electrode surface. We demonstrate here that we can affect the conformation of IgGs, likely pushing/pulling electrostatically Fab fragments towards/from the electrode surface. A potential difference between electrode and solution acts on IgGs’ charged aminoacids modulating the accessibility of the specific recognition regions of Fab fragments by antigens in solution. Consequently, antibody-antigen affinity is affected by the sign of the applied potential: a positive potential enables an effective capture of antigens; a negative one pulls the fragments towards the electrode, where steric hindrance caused by neighboring molecules largely hampers the capture of antigens. Different experimental techniques (electrochemical quartz crystal microbalance, electrochemical impedance spectroscopy, fluorescence confocal microscopy and electrochemical atomic force spectroscopy) were used to evaluate binding kinetics, surface coverage, effect of the applied electric field on IgGs, and role of charged residues on the phenomenon described. These findings expand the concept of electrical control of biological reactions and can be used to gate electrically specific recognition reactions with impact in biosensors, bioactuators, smart biodevices, nanomedicine, and fundamental studies related to chemical reaction kinetics.

Multielectron systems as possible components of molecular electronics devices are attracting compelling experimental and theoretical interest. Here we studied by electrochemical scanning tunneling techniques (EC-STMicroscopy and EC-STSpectroscopy) the electron transport properties of a redox molecule endowed with two redox levels, namely, the hydroquinone/quinone (H(2)Q/Q) couple. By forming self-assembled monolayers on Au(111) of oligo-phenylene-vinylene (OPV) derivatized H(2)Q/Q moieties, we were able to explore the features of the tunneling current/overpotential relation in the EC-STS setup. The behavior of the tunneling current sheds light onto the mechanism of electron transport involving the redox levels of the H(2)Q/Q redox pair coupled to tip and substrate electrodes

The current view of the biological membrane is that in which lipids and proteins mutually interact to accomplish membrane functions. The lateral heterogeneity of the lipid bilayer can induce partitioning of membrane-associated proteins, favoring protein-protein interaction and influence signaling and trafficking. The Atomic Force Microscope allows to study the localization of membrane-associated proteins with respect to the lipid organization at the single molecule level and without the need for fluorescence staining. These features make AFM a technique of choice to study lipid/protein interactions in model systems or native membranes. Here we will review the technical aspects inherent to and the main results obtained by AFM in the study of protein partitioning in lipid domains concentrating in particular on GPI-anchored proteins, lipidated proteins, and transmembrane proteins. Whenever possible, we will also discuss the functional consequences of what has been imaged by Atomic Force Microscopy.

We show that the activity of an ion channel is correlated with the phase state of the lipid bilayer hosting the channel. By measuring unitary conductance, dwell times, and open probability of the K+ channel KcsA as a function of temperature in lipid bilayers composed of POPE and POPG in different relative proportions, we obtain that all those properties show a trend inversion when the bilayer is in the transition region between the liquid-disordered and the solid-ordered phase. These data suggest that the physical properties of the lipid bilayer influence ion channel activity likely via a fine-tuning of its conformations. In a more general interpretative framework, we suggest that other parameters such as pH, ionic strength, and the action of amphiphilic drugs can affect the physical behavior of the lipid bilayer in a fashion similar to temperature changes resulting in functional changes of transmembrane proteins.

Supported lipid bilayers composed of 1-palmitoyl-2-oleoyl-phosphatidylethanolamine (POPE) and 1-palmitoyl-2-oleoyl-phosphatidylglycerol (POPG) were assembled by the vesicle fusion technique on mica and studied by temperature-controlled atomic force microscopy. The role of different physical parameters on the main phase transition was elucidated. Both mixed (POPE/POPG 3:1) and pure POPE bilayers were studied. By increasing the ionic strength of the solution and the incubation temperature, a shift from a decoupled phase transition of the two leaflets, to a coupled transition, with domains in register, was obtained. The observed behavior points to a modulation of the substrate/bilayer and interleaflet coupling induced by the environment and preparation conditions of supported lipid bilayers. The results are discussed in view of the role of different interactions in the system. The influence of the substrate on the lipid bilayers, in terms of interleaflet coupling, can also help us in understanding the possible effect that submembrane elements like the cytoskeleton might have on the structure and dynamics of biomembranes.

We report in this paper an approach to the effective capture of IgM antibodies from antisera and solutions based on the formation of a carpet of molecules exposing thiols off a surface. Surfaces of different nature, such as OH-exposing (glass, SiO(2), metal oxides, etc.) and noble metal ones (Au, Ag, etc.), have been first functionalized in the liquid phase by suitable chemistry [3-(mercaptopropyl)trimethoxysilane or 1,4-benzenedimethanethiol]. The resulting exposed SH moieties have been further used for binding anti-A, -B, and -D IgM molecules from goat sera via a thiol exchange reaction involving the J chain and other disulfide bonds present in the IgM molecular structure. Antibodies preserve their functional activity at the surface and appear to be able to bind specifically erythrocytes of the proper group in a fast and reliable way. These results can be generalized to the use of any kind of IgM antibody and can be valuable in surface biofunctionalization in the fields of biosensors and immunoassays.

A novel approach to the preparation of ultraflat Ni substrates suitable to scanning probe microscopy imaging of immobilized polyhistidine-tagged proteins has been devised. Exploiting freshly cleaved mica, Ni thermal evaporation followed by thermal annealing in vacuum, and the template stripping method, we have obtained Ni substrates with a rms roughness as low as 0.12 nm, which bind readily polyhistidine-tagged proteins, enabling molecular resolution imaging of isolated molecules as well as of molecular submonolayers. Protein sample exposure to imidazole causes removal of the adsorbates, confirming the involvement of the polyhistidine tail in protein surface immobilization.

A model system for the electrochemical investigation of vectorial electron transfer in biological systems was designed, assembled and characterized. Gold electrodes, functionalized with a - OCH3 terminated, aromatic self-assembled monolayer, were used as a substrate for the adsorption of variants of copper-containing, redox metalloprotein azurin. The engineered azurin bears a polyhistidine tag at its C-terminus. Thanks to the presence of the solvent exposed tag, which chelates Cu2+ ions in solution, we introduced an exogenous redox centre. The different reduction potentials of the two redox centres and their positioning with respect to the surface are such that electron transfer from the exogenous copper centre and the electrode is mediated by the native azurin active site, closely paralleling electron transfer processes in naturally occurring multicentre metalloproteins. (C) 2015 Elsevier Ltd. All rights reserved.

Number of pathogenic tick-borne bacteria cause diseases in humans and animals. Wide range of conventional methods as PCR, immuno-based detection, cultivation, reverse line blot or microscopy is available for identification of bacterial pathogens. Although, these methods are used often in laboratories, they do not allow simultaneous detection of wide spectrum of bacteria. DNA chips represent technology for fast, sensitive, relatively cheap and simultaneous detection of microorganisms in biological samples. We have developed the oligo-chip for detection of different tick-borne pathogens. This method is based on detection of fluorescent signal after hybridization reaction by using laser scanner. We are also working on preparation of electronic device for detection of specific contact of two single complementary strands of DNA. This method will allow simple detection without using the fluorescent probes and expensive laser scanner. For this purpose a novel solid-state sensor for detection of bio-molecular pro...

We report on a method for covalent immobilization of 5′-thiol-modified single strand DNA probes, onto oxygen exposing surfaces by exploiting surface derivatization by 3-mercaptopropyltrimethoxysilane and subsequent intermolecular disulfide bond formation. The various steps in the formation of the molecular edifices have been characterized by X-ray photoelectron spectroscopy, quartz crystal microbalance and atomic force microscopy under liquid. Surface reaction kinetics of thiol-modified DNA probes with thiol-bearing silanes turned out to be a second-order one, possibly due to the presence of both free thiol and S–S dimers in solution. The ability of immobilized single strand DNA to bind the complementary strand has been tested and confirmed by quartz crystal microbalance measurements. The presented DNA immobilization method appears to be applicable to any surface bearing exposed hydroxyl moieties.

Extended protein monolayers were formed by multiple-step self-chemisorption and characterized by scanning force microscopy (SFM), X-ray photoelectron spectroscopy (XPS), spectroscopic ellipsometry, and cyclic voltammetry. These uniform protein monolayers were deposited on oxygen-exposing surfaces (SiO2, mica, glass, etc.) after preexposing these surfaces to 3-aminopropyltriethoxysilane (3-APTS) and reacting the silylated surfaces with glutaric dialdehyde (GD). The samples appear flat and robust under SFM imaging conditions, both at low and molecular resolution. The thickness of the three-layer structure measured by engraving the layer with the SFM tips (4.5 nm) matches fairly well the value expected for the supramolecular architecture. XPSperformed after each of the three different stages of sample preparation confirmed the presence of the elements expected (such as Cu and Sfor the azurin layer) and allowed film growth to be followed. Similar results have been obtained for the thicknesses of the different layers by spectroscopic ellipsometry. Optical absorption spectroscopy has provided data consistent with a 75% surface coverage by a protein (sub)monolayer. Redox proteins immobilized with linkers involving a similar chemistry but providing groups binding to gold (2-mercaptoethylamine and GD) rather than to oxygen-exposing surfaces made it possible to record cyclic voltammograms of single monolayers. These electrochemical experiments have confirmed retention of the protein’s redox activity upon surface immobilization. The chemisorption approach reported in the present paper appears to be applicable to all kinds of proteins. 2002 Published by Elsevier Science B.V.

This chapter starts by considering the unique properties of the “solvent of life”, that is, water. After briefly reviewing its most salient characteristics and providing information on their possible theoretical treatment, the role of water in biology will be considered, especially as far as its effect on the structure and function of biomolecules and biostructures is concerned.

The present chapter focuses on the state of the art of electrochemical experimentation on redox biomolecules both in diffusion and immobilized on an electrode’s surface. Two main classes of molecules are considered here, namely redox metalloproteins and redox cofactors. After introducing their general properties, the approaches developed for their investigation by diffusion and surface-immobilized electrochemistry are reviewed. The remainder of the chapter introduces and develops the experimental and theoretical activities of single-molecule investigation of redox metalloproteins and derivatized redox cofactors by means of the electrochemical scanning tunneling microscope up to the attainment of the holy grail of biomolecular electronics, that is, a single-protein transistor with an electrochemical gate. Its relevance for future device implementation is critically discussed.

This chapter, the last of this book, discusses possible directions along which the electrical control of biological reactions and phenomena are likely to develop in the years to come. The potential contexts encompass both biomedical and energy applications, foreseeing the involvement of both animals (humans included) and plants. The technological development of micro- and nano-electrodes and diffused power supplies (nano-batteries), along with the advances in the field of redox biology, will for sure suggest many more phenomena where direct electrochemical control is possible and advantageous in view of an always more pervasive extension of human technological control of living matter.

This chapter introduces two basic examples of direct electrical control of biomolecular conformation, namely, voltage-gated ion channels in excised, surface immobilized, biological membranes, and antibodies of the IgG type, immobilized on an electrode surface. In both cases, the action of an applied electric field results in the modulation of protein conformation mediated by the response of positively charged amino acids (lysine, arginine) moving in response to an electric field. Several concurrent pieces of evidence, coming from a number of different experimental techniques (electrophysiology measurements, EC-AFM, EIS, confocal fluorescence microscopy, electrochemical quartz crystal microbalance), prove the achievement of full control over the molecular conformation in these two very different molecules and related biological reactions, paving the way to enlarging the number of cases where the action of an applied electric field can effectively modulate biological phenomena (e.g., enzymatic reactions, etc.).

In the present chapter we will consider paradigmatic cases in which the role of the redox state of certain molecular effectors (e.g., enzymes, cofactors, etc.) has been shown to be a determinant in modulating the gene expression level through a cascade of molecular interactions and events. We will consider only those cases in which the redox functionality has been shown in a robust, unequivocal way. These examples will serve as a starting point for the remainder of the book, where a novel approach will be suggested, based on direct electrochemistry, to influence those redox pathways in order to drive the phenomenon of gene expression at a technological level and, possibly, to alter in a predictable fashion the fate of a given organism. We will focus on photosynthetic bacteria, E. coli and sub-cellular organelles such as chloroplasts and mitochondria, whose gene regulation is partially or fully redox dependent.

We review the capabilities of Atomic Force Microscopy (AFM) in the study of phase transitions in Supported Lipid Bilayers (SLBs). AFM represents a powerful technique to cover the resolution range not available to fluorescence imaging techniques and where spectroscopic data suggest what the relevant lateral scale for domain formation might be. Phase transitions of lipid bilayers involve the formation of domains characterized by different heights with respect to the surrounding phase and are therefore easily identified by AFM in liquid solution once the bilayer is confined to a flat surface. Even if not endowed with high time resolution, AFM allows light to be shed on some aspects related to lipid phase transitions in the case of both a single lipid component and lipid mixtures containing sterols also. We discuss here the obtained results in light of the peculiarities of supported lipid bilayer model systems. © the Partner Organisations 2014.

This chapter provides the reader with some basic notions in electrochemistry that will serve to form the language and the interpretation framework for all of the scientific results and ideas that will be presented in the following chapters. Starting from the basic physical-chemistry of a charged surface (e.g., an electrode) in a water-based solution, the chapter will introduce the concept of electrochemical reactions at electrodes, focusing on the fundamental microscopic theory of electron transfer by Marcus. This section will be followed by one introducing basic electrochemical tools for driving the potential of one or two working electrodes in solution and by a review of the electrochemical techniques that are used in the book, along with some key theory for data interpretation.

The redox metalloprotein azurin self-chemisorbed onto Au(1 1 1) substrates has been investigated by electrochemically controlled scanning tunneling (STM) and scanning force/lateral force microscopy (SFM/LFM) and cyclic voltammetry (CV) in aqueous solution. The combined use of STM and SFM/LFM under electrochemical control in the negative side of the azurin redox midpoint (+116 mV vs. SCE) has delivered unique information on the nature of the STM images. While in STM the bright spots, believed to be associated with azurin molecules, are visible only for potential values higher than −125 mV, the concurrent electrochemical SFM results show adsorbed proteins over the whole potential range investigated (from −225 to +75 mV). Stepping the potential back and forth (between −25 and −125 mV) in STM imaging, it has been possible to make bright spots appearing and disappearing repeatedly, indicating that STM image formation arises possibly through resonant tunneling via the redox levels of azurin. These results represent the first clear evidence of potential-dependent tunneling in proteins adsorbed onto a conductive substrate.

The role of chain length in affecting the molecular organization in thin films of mesogenic Zn−porphyrin octaesters ZnCnOAP has been investigated by means of Langmuir isotherms and area-per-molecule relaxation measurements at the air−water interface, as well as by unpolarized and polarized optical absorption spectroscopy and polarized ATR-FTIR spectroscopy after transfer onto the solid substrate. The results clearly identify three different classes of behavior depending on the chain length. Intermediate chain length molecules (ZnCnOAP, where n = 6−12) feature clear coexistence regions in the isotherms as well as marked molecular relaxation due to self-organization effects in the coexistence region. Such behavior is absent in the short-chain derivative ZnC3OAP and strongly reduced in the longest side chain derivative ZnC14OAP, reflecting the tendency of intermediate chain length porphyrins to form columnar mesophases in the bulk. These data are confirmed by the time evolution of the optical absorption spec...

We have performed inelastic and quasielastic neutron-scattering experiments (IQENS) on the highly lithium doped fulleride ${\mathrm{Li}}_{12}{\mathrm{C}}_{60}$ covering the T range from 150 to 600 K, across a tetragonal to cubic fcc phase transition which was shown to occur around $T=550\mathrm{K}$ by synchrotron x-ray diffraction. In this work we focus on the dynamics of the Li ions and ${\mathrm{C}}_{60}$ molecular units. The QENS data suggest a dynamical model in which the Li ions diffuse in the confined environment of the octahedral void of the pristine ${\mathrm{C}}_{60}$ molecule, in agreement with the structural information that we have obtained on the same compound. These results are discussed together with our previously reported analysis of the changes in the vibrational density of states across the phase transition. The emerging picture is that ${\mathrm{Li}}_{12}{\mathrm{C}}_{60}$ at high temperature (fcc phase) has the same dynamical features as pure ${\mathrm{C}}_{60}$ with the addition of Li localized diffusion within the octahedral void, while the low-temperature tetragonal phase is characterized by a broadening and disappearance of many dynamical features, i.e., of the Li diffusion and of the ${\mathrm{C}}_{60}$ librations and molecular modes.

We present results on the dynamics of the Li+ ions and of the C60 molecular units in Li12C60 obtained by INS and QENS experiments performed on IN6 (at the Institut Laue-Langevin) covering the temperature range 150K to 600 K. The transition from the high temperature Fm3m to the Iow temperatur tetragonal structure of Li12C60 is accompanied by significant changes in the vibrational density of states (v-DOS). The role of the Li+ vibrational and diffusional motion in determining the changes in the overall v-DOS is investigated.

The time dependence of the adsorption of myelin basic protein onto dipalmitoyl phosphatidyl glycerol multilayers has been followed directly, using a novel application of a microgravimetric gauge. Our results, supplemented by other data obtained by FTIR, show the ease and versatility of the quartz microbalance for investigating the interaction processes between proteins and phospholipid layers and show that the protein adsorption is accompanied by structural changes in the proteolipid ensemble and adsorbed liquid water; it is furthermore dependent on the mesoscopic defect morphology of the ensemble.

We have measured the relaxation to equilibrium in the supercooled nematic phase upon approach to of a fragile liquid-crystalline photosensitive polymeric glass former. Perturbation is introduced in a non-conventional way by optical pumping (orientation) of the nematic polydomain phase. Relaxation back to equilibrium is followed both on the microscopic scale, by means of depolarized time-resolved Raman experiments, and on the macroscopic scale, by means of quartz crystal microgravimetry.

Ultrathin semiconductor layers made of CdS nanocrystals synthesized in cadmium arachidate Langmuir-Blodgett films have been investigated by means of polarized optical and infrared absorption spectroscopy, resonance Raman scattering and X-ray diffraction during two different phases of the formation process (before and after the removal of the organic precursor matrix). Optical spectroscopy revealed that CdS nanocrystals are anisotropic, likely ellipsoidal in shape, laying with their major axes in the film plane. Resonance Raman scattering confirmed these results by monitoring, in two different experimental geometries and after different annealing conditions, the coupling to the lattice of the lowest electronic excited state. The reported data allow to describe these samples as made of anisotropic CdS nanocrystals surrounded by a small residual amount of fatty acids. Finally, we have demonstrated the possibility of varying the CdS crystal size from 3 to 10 nm upon the removal of the organic matrix and the annealing procedure.

The morphology of ultrathin layers of CdS nanocrystals from cadmium arachidate Langmuir–Blodgett (LB) film templates has been investigated by means of scanning force and scanning electron microscopies (SEM). Consistently with previous Brewster angle microscopy investigations, low resolution electron micrographs reveal a flat film surface characterized by steps due to the difference in the number of precursor layers. Scanning force microscopy (SFM) allows one to get information on the mesoscopic and atomic film structure. While in the range 1–10 μ m the film appears to be formed by a series of flat platforms, often shaped like triangles and sometimes superorganized in larger needle-like structures, in the hundred nanometer scale an irregular granular structure appears. Finally, performing zooms onto flat platforms, the CdS hexagonal lattice appears, revealing a lattice periodicity of 0.42 nm, in agreement with the bulk value.

The structural modifications of photosynthetic reaction centers from Rhodobacter sphaeroides arranged in Langmuir films and after deposition onto solid substrate have been investigated as well as the variations in protein orientation as a function of surface pressure. While at low surface pressure (15-25 mN/m) protein integrity appears to be affected by the action of the surface tension, as results from the film behavior at the air-water interface and from optical spectroscopy data. When the surface pressure (30-45 mN/m) is increased, protein structure is much better preserved. Moreover, at those deposition pressures, the film is characterized not only by a higher surface density but also by an increased anisotropy, as revealed by polarized absorption spectroscopy and confirmed by optical ellipsometry. Measurements of water content desorption in deposited films by means of a gravimetric transducer have consistently confirmed the preservation of protein structure at higher surface pressures.

An integrated field-effect device for fully electronic deoxyribonucleic acid (DNA) detection was realized in a standard CMOS process. The device is composed of a floating-gate MOS transistor, a control-capacitor acting as integrated counterelectrode, and an exposed active area for DNA immobilization. The drain-current of the transistor is modulated by the electric charge carried by the DNA molecules. After DNA hybridization, this charge increases and a change in the output current is measured. Experimental results are provided. Full compatibility with a standard CMOS process-opens the way to the realization of low-cost large-scale integration of fast electronic DNA detectors.

A novel method for the formation of ultrathin polycrystal semiconductor layers based on the Langmuir-Blodgett technique is reported. The proposed approach allows the synthesis of layers of pure compounds belonging to the II–VI groups such as CdS and PbS with a thickness resolution of 0·8 nm starting from Langmuir-Blodgett films of fatty acid salts exposed to an atmosphere of hydrogen sulfide. The layers formed are characterized by Brewster angle microscopy on the solid substrate showing good uniformity and constant thickness. This new method could have interesting applications in the formation of thin sensitive layer transducers for enhanced performance field-effect based biosensors such as ChemFET or ISFET.

Nanometer size semiconductor (PbS) particles were synthesized in the fatty acid matrix by exposing the fatty acid salt Langmuir-Blodgett films to an atmosphere of H 2 S. Realization of the structure: electrode-tunnelling gap-particle-gap-STM tip allowed one to register monoelectron phenomena, such as Coulomb blockade and Coulomb staircase. Synthesis of the nanoparticle at the tip of a metal stylus allowed one to avoid the necessity of using STM and permitted one to register monoelectron characteristics using only a feedback in one direction perpendicular to the flat electrode plane. Another phenomenon connected with negative resistance regions in voltage/current characteristics was observed just by varying the tip-flat electrode distance. It was found that bulk properties of the particle material do not play a crucial role in the appeatance of the monoelectron properties.

The technique of electrochemical scanning tunneling microscopy (ECSTM) and spectroscopy (ECSTS) for studying electron transport through single redox molecules is here described. Redox molecules of both biological and organic nature have been studied by this technique with the aim of understanding the transport mechanisms ruling the flow of electrons via a single molecule placed in a nanometer-sized gap between two electrodes while elucidating the role of the redox density of states brought about by the molecule. The obtained results provide unique clues to single-molecule transport behavior and support the concept of single-molecule electrochemical gating.

The technique of electrochemical scanning tunneling microscopy (ECSTM) and spectroscopy (ECSTS) for studying electron transport through single redox molecules is here described. Redox molecules of both biological and organic nature have been studied by this technique with the aim of understanding the transport mechanisms ruling the flow of electrons via a single molecule placed in a nanometer-sized gap between two electrodes while elucidating the role of the redox density of states brought about by the molecule. The obtained results provide unique clues to single-molecule transport behavior and support the concept of single-molecule electrochemical gating.

The possibility of controlling electron transport in a single molecule bridged between two metal electrodes represents the ultimate goal of molecular electronics. Molecular electronics aims also at introducing specific properties for the electron transport features both by controlling the structural details of the junction and by exploiting new chemical functionalities. Here we show that, in a molecular junction, where electrodes are represented by a gold substrate and the tip of a scanning tunneling microscope in electrochemical environment, the use of a single molecular species makes it possible to obtain different features for the tunneling current according to the structural details of the junction. In particular, molecules endowed with redox properties brought about by a hydroquinone/benzoquinone redox couple can show both transistor-like and negative differential resistance (NDR) effects. We discuss the mechanistic processes that might describe the different behavior in light of theories of electron transfer between metal electrodes and redox molecules. The results show, on the one hand, the great potential and flexibility that molecular electronics offer and, on the other hand, the need of controlling as much as possible the details of the tunneling junction in order to obtain reproducible results.