The association between real-life markers of phone use and cognitive performance, health-related quality of life and sleep.

INTRODUCTION: The real-life short-term implications of electromagnetic fields (RF-EMF) on cognitive performance and health-related quality of life have not been well studied. The SPUTNIC study (Study Panel on Upcoming Technologies to study Non-Ionizing radiation and Cognition) aimed to investigate possible correlations between mobile phone radiation and human health, including cognition, health-related quality of life and sleep. METHODS: Adult participants tracked various daily markers of RF-EMF exposures (cordless calls, mobile calls, and mobile screen time 4 h prior to each assessment) as well as three health outcomes over ten study days: 1) cognitive performance, 2) health-related quality of life (HRQoL), and 3) sleep duration and quality. Cognitive performance was measured through six "game-like" tests, assessing verbal and visuo-spatial performance repeatedly. HRQoL was assessed as fatigue, mood and stress on a Likert-scale (1-10). Sleep duration and efficiency was measured using activity trackers. We fitted mixed models with random intercepts per participant on cognitive, HRQoL and sleep scores. Possible time-varying confounders were assessed at daily intervals by questionnaire and used for model adjustment. RESULTS: A total of 121 participants ultimately took part in the SPUTNIC study, including 63 from Besancon and 58 from Basel. Self-reported wireless phone use and screen time were sporadically associated with visuo-spatial and verbal cognitive performance, compatible with chance findings. We found a small but robust significant increase in stress 0.03 (0.00-0.06; on a 1-10 Likert-scale) in relation to a 10-min increase in mobile phone screen time. Sleep duration and quality were not associated with either cordless or mobile phone calls, or with screen time. DISCUSSION: The study did not find associations between short-term RF-EMF markers and cognitive performance, HRQoL, or sleep duration and quality. The most consistent finding was increased stress in relation to more screen time, but no association with cordless or mobile phone call time.

Orthogonal Chemical Functionalization of Au/SiO2/TiW Patterned Substrates.

Macropatterned and micropatterned gold/silicon dioxide/titanium tungsten (Au/SiO2/TiW) substrates were orthogonally functionalized: three different molecules (monovalent silane, thiol, and phosphonic acid) were used to specifically form organolayers on Au, SiO2, or TiW areas of patterned substrates. The orthogonality of the functionalization (i.e., selective grafting of thiol on Au, phosphonic acid on TiW, and silane on SiO2) was assessed by X-ray photoelectron spectroscopy (XPS), time-of-flight secondary ion mass spectrometry (ToF-SIMS), Fourier transform infrared spectroscopy (FTIR), and contact angle measurements. These results are especially promising for the selective anchoring of targets (e.g., biomolecules, nanoparticles, nanowires, nanotubes, or other nano-objects) onto patterned zones of multimaterial substrates, such as nanosensors or other nanodevices.

Oxidized Titanium Tungsten Surface Functionalization by Silane-, Phosphonic Acid-, or Ortho-dihydroxyaryl-Based Organolayers.

Titanium tungsten (TiW) films (200 nm thick) were cleaned by oxygen plasma, and the resulting oxidized surfaces were functionalized by 3-aminopropylphosphonic acid (APPA), 3-ethoxydimethylsilylpropylamine (APDMES), or dopamine (DA) to form three different organolayers. The three resulting organolayers were characterized by X-ray photoelectron spectroscopy, time-of-flight secondary ion mass spectrometry, and Fourier transform infrared spectroscopy analyses. The stability of each organolayer was investigated. Our results suggested that the Si-O-Ti or Si-O-W bonds formed by the reactions of APDMES with surface-oxidized TiW were rather labile, whereas the catechol layer was less labile. The APPA layer was the most stable of all tested surface modifications.

Orthogonal chemical functionalization of patterned Au/TiW substrate for selective immobilization of nanoparticles.

The evolution of nanobiosensors stresses the need for multi-material nanopatterned surfaces to enhance sensing performances. Titanium tungsten (TiW) has been mastered and routinely implemented in nanoelectronic devices, in a reproducible way and at industrial production scales. Such a material may be envisioned for use in (bio)chemical nanoelectronic sensors, but the surface functionalization of such material has yet to be studied. In the present article, the orthogonal chemical functionalization of patterned Au on TiW substrates has been explored for the first time. Surface functionalizations were assessed by x-ray photoelectron spectroscopy, polarization modulation infrared reflection-absorption spectroscopy and time-of-flight secondary ion mass spectrometry imaging. Au/TiW patterned substrates were functionalized with mercapto-undecamine. Thanks to the orthogonality of thiol/Au versus phosphonic acid/TiW reactions, only the Au features were modified leading to the amine derivatized surface. This allowed for the localizing of carboxy-functionalized nanoparticles by electrostatic interaction on Au with a selectivity above 10 when compared to TiW.

Screening of a Library of Oligosaccharides Targeting Lectin LecB of Pseudomonas Aeruginosa and Synthesis of High Affinity Oligoglycoclusters.

The Gram negative bacterium Pseudomonas aeruginosa (PA) is an opportunistic bacterium that causes severe and chronic infection of immune-depressed patients. It has the ability to form a biofilm that gives a selective advantage to the bacteria with respect to antibiotherapy and host defenses. Herein, we have focused on the tetrameric soluble lectin which is involved in bacterium adherence to host cells, biofilm formation, and cytotoxicity. It binds to l-fucose, d-mannose and glycan exposing terminal fucose or mannose. Using a competitive assay on microarray, 156 oligosaccharides and polysaccharides issued from fermentation or from the biomass were screened toward their affinity to LecB. Next, the five best ligands (Lewisa, Lewisb, Lewisx, siayl-Lewisx and 3-fucosyllactose) were derivatized with a propargyl aglycon allowing the synthesis of 25 trivalent, 25 tetravalent and 5 monovalent constructions thanks to copper catalyzed azide alkyne cycloaddition. The 55 clusters were immobilized by DNA Directed immobilization leading to the fabrication of a glycocluster microarray. Their binding to LecB was studied. Multivalency improved the binding to LecB. The binding structure relationship of the clusters is mainly influenced by the carbohydrate residues. Molecular simulations indicated that the simultaneous contact of both binding sites of monomer A and D seems to be energetically possible.

Design and Synthesis of Galactosylated Bifurcated Ligands with Nanomolar Affinity for Lectin LecA from Pseudomonas aeruginosa.

Lectin A (LecA) from Pseudomonas aeruginosa is an established virulence factor. Glycoclusters that target LecA and are able to compete with human glycoconjugates present on epithelial cells are promising candidates to treat P. aeruginosa infection. A family of 32 glycodendrimers of generation 0 and 1 based on a bifurcated bis-galactoside motif have been designed to interact with LecA. The influences both of the central multivalent core and of the aglycon of these glycodendrimers on their affinity toward LecA have been evaluated by use of a microarray technique, both qualitatively for rapid screening of the binding properties and also quantitatively (Kd ). This has led to high-affinity LecA ligands with Kd values in the low nanomolar range (Kd =22 nm for the best one).

Toward the Rational Design of Galactosylated Glycoclusters That Target Pseudomonas aeruginosa Lectin A (LecA): Influence of Linker Arms That Lead to Low-Nanomolar Multivalent Ligands.

Anti-infectious strategies against pathogen infections can be achieved through antiadhesive strategies by using multivalent ligands of bacterial virulence factors. LecA and LecB are lectins of Pseudomonas aeruginosa implicated in biofilm formation. A series of 27 LecA-targeting glycoclusters have been synthesized. Nine aromatic galactose aglycons were investigated with three different linker arms that connect the central mannopyranoside core. A low-nanomolar (Kd =19 nm, microarray) ligand with a tyrosine-based linker arm could be identified in a structure-activity relationship study. Molecular modeling of the glycoclusters bound to the lectin tetramer was also used to rationalize the binding properties observed.

Anti-heat shock protein autoantibody profiling in breast cancer using customized protein microarray.

Heat shock proteins (HSPs) are over-expressed in a wide range of human cancers. It results in the stimulation of the immune system and consequently in elevated concentration of anti-HSP autoantibodies. Elevated anti-HSP autoantibodies were found in breast cancer patients, and they are associated with tumor metastasis. Therefore, screening these autoantibodies could be of diagnostic and prognostic values. Protein microarrays have already demonstrated their great potential as a diagnostic tool. However, protein diversity requires optimization of the microarray fabrication to achieve high sensitivity and specificity. In this study, seven HSPs were immobilized on six different surface chemistries. After evaluation and optimization with purified antibodies of the six surface chemistries, two surfaces were selected to detect anti-HSP autoantibodies in breast cancer sera. Multiplex detection of anti-HSP autoantibodies allowed discrimination of breast cancer patients (50) from healthy controls (26) with a sensitivity of 86% and a specificity of 100%.

Effects of the Surface Densities of Glycoclusters on the Determination of Their IC50 and Kd Value Determination by Using a Microarray.

Pseudomonas aeruginosa (PA) is an opportunistic bacterium involved in 10-30% of nosocomial diseases. It causes severe lung injury to cystic fibrosis patients, often leading to patient death. PA strains are multidrug resistant, thus making the design of new therapeutics a challenge for public health. One promising therapeutic option is to design glycoclusters that target the virulence factor of PA. LecA is a galactose-specific lectin that might be involved in adhesion and biofilm formation by PA. The DNA-directed immobilization (DDI) microarray is a powerful tool for screening and understanding of structure-activity relationships between glycoclusters and lectins. High-throughput and multiplexed analysis of lectin-glycocluster interactions on a DDI microarray allows measurement of IC50 and dissociation constant (Kd ) values with minute amounts of material. In order to study the robustness of the DDI microarray in determination of IC50 and Kd values, the impact of glycocluster surface density was investigated. The data obtained show that measured IC50 values were influenced by glycocluster surface density: as the density of glycoclusters increases, the measured IC50 values increase too. In contrast, the measured Kd values were not affected by glycocluster surface density, provided that the experimental conditions allow interaction between glycocluster and lectin at single-molecule level (no surface cluster effect).

Structure binding relationship of galactosylated Glycoclusters toward Pseudomonas aeruginosa lectin LecA using a DNA-based carbohydrate microarray.

Pseudomonas aeruginosa (PA) is a major public health issue due to its impact on nosocomial infections as well as its impact on cystic fibrosis patient mortality. One of the main concerns is its ability to develop antibiotic resistance. Therefore, inhibition of PA virulence has been proposed as an alternative strategy to tackle PA based infections. LecA (or PA-IL), a galactose binding lectin from PA, is involved in its virulence. Herein, we aimed at designing high affinity synthetic ligands toward LecA for its inhibition and at understanding the key parameters governing the binding of multivalent galactosylated clusters. Twenty-five glycoclusters were synthesized and their bindings were studied on a carbohydrate microarray. Monosaccharide centered clusters and linear comb-like clusters were synthesized with different linkers separating the core and the galactosyl residues. Their length, flexibility, and aromaticity were varied. Our results showed that the binding profile of LecA to galactosylated clusters was dependent on both the core and the linker and also that the optimal linker was different for each core. Nevertheless, an aryl group in the linker structure drastically improved the binding to LecA. Our results also suggest that optimal distances are preferred between the core and the aromatic group and the core and the galactose.

The influence of the aromatic aglycon of galactoclusters on the binding of LecA: a case study with O-phenyl, S-phenyl, O-benzyl, S-benzyl, O-biphenyl and O-naphthyl aglycons.

A library of 24 new mannose-centered tetragalactoclusters with four different linkers (di- and triethyleneglycol with phosphodiester or phosphorothioate linkages) and six different aromatic aglycons (O-phenyl, S-phenyl, O-benzyl, S-benzyl, O-biphenyl and O-naphthyl) was synthesized. Their interactions with LecA were evaluated on a DNA Directed Immobilization (DDI) based glycocluster array allowing the determination of their IC50 against lactose and the evaluation of their dissociation constant (Kd). Finally, the docking simulations confirm the experimental results and demonstrated that the better affinity of O-biphenyl- and O-naphthyl-galactoside is due to a double interaction between the aromatic ring and the histidine 50 and proline 51 of LecA.

Characterization of three amino-functionalized surfaces and evaluation of antibody immobilization for the multiplex detection of tumor markers involved in colorectal cancer.

Antibody microarrays are powerful and high-throughput tools for screening and identifying tumor markers from small sample volumes of only a few microliters. Optimization of surface chemistry and spotting conditions are crucial parameters to enhance antibodies' immobilization efficiency and to maintain their biological activity. Here, we report the implementation of an antibody microarray for the detection of tumor markers involved in colorectal cancer. Three-dimensional microstructured glass slides were functionalized with three different aminated molecules ((3-aminopropyl)dimethylethoxysilane (APDMES), Jeffamine, and chitosan) varying in their chain length, their amine density, and their hydrophilic/hydrophobic balance. The physicochemical properties of the resulting surfaces were characterized. Antibody immobilization efficiency through physical interaction was studied as a function of surface properties as well as a function of the immobilization conditions. The results show that surface energy, steric hindrance, and pH of spotting buffer have great effects on protein immobilization. Under optimal conditions, biological activities of four immobilized antitumor marker antibodies were evaluated in multiplex immunoassay for the detection of the corresponding tumor markers. Results indicated that the chitosan functionalized surface displayed the highest binding capacity and allowed to retain maximal biological activity of the four tested antibody/antigen systems. Thus, we successfully demonstrated the application of amino-based surface modification for antibody microarrays to efficiently detect tumor markers.

Effects of Silane Monolayers on Lysophosphatidylcholine (LysoPC) Detection by Desorption Ionization on Silicon Mass Spectrometry (DIOS-MS) in Solution and Plasma.

Desorption ionization on silicon mass spectrometry (DIOS-MS) enables high throughput analysis of low-molecular-weight biomolecules. However, detection of metabolite biomarkers in complex fluids such as plasma requires sample pretreatment, limiting clinical application. Here, we show that porous silicon, chemically modified using monolayers of n-propyldimethylmethoxysilane molecules, is a good candidate for fingerprinting lysophosphatidylcholine (lysoPC) in plasma, without sample pretreatment, for DIOS-MS-based diagnosis (e.g., sepsis). Results were correlated to lysoPC molecule location inside/outside the pores, determined by time-of-flight secondary ion mass spectrometry profiling, and to physicochemical properties.

Effect of Silane Monolayers and Nanoporous Silicon Surfaces on the Matrix-Assisted Laser Desorption Ionization Mass Spectrometry Detection of Sepsis Metabolites Biomarkers Mixed in Solution.

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-ToF MS) is a promising strategy for clinical diagnosis based on metabolite detection. However, several bottlenecks (such as the lack of reproducibility in analysis, the presence of an important background in low-mass range, and the lack of organic matrix for some molecules) prevent its transfer to clinical cases. These limitations can be addressed by using nanoporous silicon surfaces chemically functionalized with silane monolayers. In the present study, sepsis metabolite biomarkers were used to investigate the effects of silane monolayers and porous silicon substrates on MALDI-ToF MS analysis (signal-to-noise value (S/N), relative standard deviation of the S/N of triplicate samples (STDmean), and intra-substrates uniformity). Also, the impact of the physicochemical properties of metabolites, with different isoelectric points and hydrophobic-hydrophilic balances, was assessed. Four different silane molecules, with various alkyl chain lengths and head-group charges, were self-assembled in monolayers on plane and porous silicon surfaces. Their surface coverage and conformity were investigated by X-ray photoelectron spectroscopy (XPS) and time-of-flight secondary ion mass spectrometry (ToF-SIMS). The seven metabolites detected on the stainless-steel target plate (lysophosphatidylcholine, caffeine, phenylalanine, creatinine, valine, arginine, and glycerophosphocholine) are also detected on the silanized and bare, plane and porous silicon surfaces. Moreover, two metabolites, glycine and alanine, which are not detected on the stainless-steel target plate, are detected on all silanized surfaces, except glycine which is not detected on CH3 short-modified porous silicon and on the bare plane silicon substrate. In addition, whatever the metabolites (except phenylalanine and valine), at least one of the silicon surfaces allows to increase the S/N value in comparison with the stainless-steel target plate. Also, the heterogeneity of matrix crystallization features is linked to the STDmean which is poor on the NH3+ monolayer on plane substrate and better on the NH3+ monolayer on porous substrate, for most of the metabolites. Nevertheless, matrix crystallization features are not sufficient to systematically get high STDmean and uniformity in MALDI-ToF MS analysis. Indeed, the physicochemical properties of metabolites and surfaces, limitations in metabolite extraction from the pores, and improvement in metabolite desorption due to the pores are shown to significantly impact MS analysis. In particular, in the case of the most hydrophobic metabolites studied, the highest S/N values and the best STDmean and uniformity (the lowest values) are reached by using porous substrates, while in the case of the most hydrophilic metabolites studied, plane substrates demonstrated the highest S/N and the lowest STDmean. No clear trend of surface chemistry was evidenced.

Nanoparticles selectively immobilized onto large arrays of gold micro and nanostructures through surface chemical functionalizations.

Latex nanoparticles (100nm and 200nm diameter) were precisely located onto the gold regions of micro and nanopatterned gold/silica substrates through surface chemical functionalizations. The gold patterns were selectively functionalized with alkylthiols bearing biotin or amine headgroups. This selective functionalization allowed the trapping of streptavidin- or carboxy-functionalized latex nanoparticles onto the gold structures with very little non-specific adsorption onto the surrounding silica. Quantitative data of nanoparticle capture on gold and silica, obtained through SEM image analysis, showed a one to two order of magnitude increase on gold with a similar low coverage on silica (non-specific adsorption) thanks to chemical functionalizations. Single nanoparticles were captured at the gap of dimer gold nanostructures.

DNA directed immobilization glycocluster array: applications and perspectives.

The present review concerns the recent advances in DNA directed immobilization (DDI) based glycocluster array. The impact of glycan immobilization on subsequent interactions with protein is discussed and the consequent pros and cons of DDI-based glycocluster array are reviewed. Finally, application in the discovery of anti-pathogen molecules is illustrated by screening for galactose or fucose glycoclusters targeting two Pseudomonas aeruginosa virulence factors (PA-IL and PA-IIL).

Improvement of protein immobilization for the elaboration of tumor-associated antigen microarrays: application to the sensitive and specific detection of tumor markers from breast cancer sera.

There is an urgent need to identify relevant tumor markers showing high sensitivity and specificity for early diagnosis and prognosis of breast cancer. Protein microarrays have demonstrated to be cost-effective, high through-put and powerful tools for screening and identifying tumor markers with only minute samples. Autoantibodies directed against tumor-associated antigens (TAAs) were shown to be relevant tumor markers. However, due to the variability of immune response from one individual to another and depending on the type of cancer, detection of only one type of anti-TAA autoantibody is not sufficient to give a reliable and precise diagnosis. It is necessary to use a set of several TAAs for determining specific autoimmune profiles. Therefore, combining various TAAs on different surfaces could improve sensitivity and specificity for anti-TAA autoantibody detection. Herein a panel of 10 proteins, including well-known tumor-associated antigens (TAAs) and potential new biomarkers of breast cancer, were immobilized onto microstructured microarray under optimized conditions (spotting pH buffer, surface chemistry, blocking procedure), in order to determine an autoimmune signature of breast cancer. Sera from 29 breast cancer patients and 28 healthy donors were screened in sandwich immunoassays on the miniaturized system to detect the eventual presence of anti-TAAs autoantibodies. Results indicated that the detection level of each anti-TAA autoantibody in a given serum sample was strongly dependant on the surface chemistry. Combining five TAAs (p53, Hsp60, Hsp70, Her2-Fc, NY-ESO-1) on two different surface chemistries (NHS and APDMES) allowed the significant detection of more than 82% breast cancer sera.

Quantitative analysis (K(d) and IC(50)) of glycoconjugates interactions with a bacterial lectin on a carbohydrate microarray with DNA Direct Immobilization (DDI).

Nowadays, there is a great interest for understanding the structure/function relationship governing recognition of carbohydrates by their receptors for the design of new treatments. Indeed, carbohydrates and glycoconjugates play a major role in key biological events such as cell-cell recognition, pathogenesis inflammation, and host pathogen interactions. Pseudomonas aeruginosa (PA) is one of the predominant bacterium encountered in nosocomial infections. PA infections often lead to chronic inflammation and eventually to death despite aggressive antibiotic therapy: the emergence of resistant strains and biofilm formation seems to give a selective advantage to the bacterium. A promising approach is to inhibit the virulence factors of PA such as PA-IL which is a galactose specific lectin. Herein, we develop a microarray to probe the binding of six galacto-conjugates to PA-IL differing by their spatial configuration and geometry. This microsystem is made of 40 independent microwells in which 64 spots of glycoconjugates probes are arrayed by using DNA Directed Immobilization (DDI). This microsystem allows, in a multiplex fashion, qualitative information on the binding by direct fluorescence readout as well as quantitative information by the determination of IC(50) values in a competition assay and surface dissociation constants (K(d)). According to our data, direct fluorescent signals (FI(635)), IC(50) and K(d) values provided similar ranking for glycoconjugates with respect to PA-IL binding thus affording a general tool for the selection of galacto-conjugates displaying the best affinities toward PA-IL.