Yanxia Hou

Quantitative determination of zinc in milkvetch by anodic stripping voltammetry with bismuth film electrodes

Bismuth film electrode (BiFE) was shown to be an attractive alternative to common mercury film electrode (MFE) for anodic stripping voltammetric measurements. In this study, bismuth film, that was in situ deposited onto glassy carbon electrode, was used to detect zinc content of milkvetch, used in traditional Chinese medicine. Variables affecting the response have been evaluated and optimized. Experimental results showed a high response, with a good linearity (between 0.5x10(-6)molL(-1) and 3x10(-6)molL(-1)) a good precision (R.S.D.=3.58%) and a low detection limit (9.6x10(-9)molL(-1) with a 120s anodic). The anodic stripping performance makes the bismuth film electrode very desirable for measurements of trace nutritive element zinc in milkvetch and should impart possible restrictions on the use of mercury electrode.

Mixed urease/amphiphile LB films and their application for biosensor development

The optimal conditions for the formation of a stable mixed urease/octadecylamine (ODA) film at the air-water interface are determined. This film is efficiently transferred onto the hydrophobized surface of a pH-ISFET, and the features of the urea enzymatic field-effect transistor (ENFET) are determined: detection limit 0.2 mM, response time 15 s, and dynamic range 0-20 mM.

Optimization of the mixed urease/amphiphile Langmuir–Blodgett film and its application for biosensor development

The interaction of urease in subphase at a certain pH value with amphiphiles at the air-water interface leading to the formation of a stable mixed monolayer is of special interest due to its potential application in biosensor development. The dependence of stability of the mixed monolayers on various factors such as amphiphilic type (octadecylamine and behenic acid in this case), urease concentration in subphase, adsorption time, surface pressure and temperature of the subphase are studied in this paper. Octadecylamine, as amphiphilic molecule, shows a stronger associating ability with urease than behenic acid when urease concentration is kept in a suitable range of 4-12 mg/l. Mixed urease/ octadecylamine monolayer can efficiently be transferred onto the hydrophobic surface of octadecylsilane-modified SiO 2 /Si substrate with an ideal transfer ratio. This is the crucial point to meet requirements for a reproducible deposition on a transducer surface, such as pH-ISFET (ion-sensitive field effect transistors), to achieve good biosensor characteristics.

Continuous Evolution Profiles for Electronic‐Tongue‐Based Analysis

Conventional biosensors and biochips rely on a “lock-andkey” recognition principle, for which specific ligands, such as aptamers, antibodies, or mimetics are needed. In contrast, electronic nose/tongue devices (eN/eT) use the differential binding of analytes, either alone or in mixtures, to an array of cross-reactive receptors (CRRs) whose combined responses create a characteristic pattern for each component. As a result, none of the CRRs need to be highly specific for any given analyte and the time-consuming design and generation of specific sensors may be circumvented. Based on this principle, CRR arrays associated with appropriate detection have been developed. For example, synthetic tetraphenylporphyrin derivatives, whose fluorescence quenching upon protein binding was used for detection, or a receptor library, integrating a peptide cavity coupled with indicator-uptake colorimetric detection, were developed for protein sensing above concentrations of ten micromolar. A sensor array composed of gold nanoparticles, grafted with different amino-functionalized thiols and conjugated with an anionic fluorescent polymer (poly(p-phenyleneethynylene, PPE), allowed detection of proteins in the low nanomolar range, as well as detection of bacteria or discrimination of normal cells from their cancerous counterparts. Furthermore, replacing PPE with green fluorescent protein led to a concentration-sensitive CRR array able to discriminate different proteins in human serum at physiologically relevant concentrations. However, though simplified with regard to conventional biosensors, the preparation of eN/eT sensing arrays still requires the design and synthesis 5–29 CRRs and conjugation to a ligand-independent detection system. The development of new eN/eTapplications would thus benefit from simplified production of the sensing elements. In this regard, eN applications take advantage of the differential diffusion of volatile biomarkers through films of a single sensing material in varying morphology and surface coverage. For an eT, we anticipated that the design and synthetic efforts could also be drastically reduced if the CRRs were prepared by selfassembly of different combinations of building blocks (BBs), that is easily accessible molecules displaying different physicochemical properties. In this way, simply mixing different BBs in varying and controlled proportions and allowing them to assemble, either alone or on a template, should lead to a collection of combinatorial cross-reactive receptors (CoCRRs) with evolutionary properties. Interestingly, a high diversity of CoCRRs can be produced from a restricted number of BBs: 11 with only two BBs mixed in concentrations varying from 0 to 100% in 10% increments and 66 by simply adding a third BB. Further generalization to n BBs and i% concentration increment leads to [(100/i)+ n 1]!/[(n 1)!(100/i)!] potential different CoCRRs. To assure 1) reproducible assembly; 2) close correlation between the CoCRRs topology and the precursor combination of BBs; 3) spatial encoding of the CoCRRs compositions, we reasoned that the BBs would best be combined through the formation of self-assembled monolayers (SAMs). Interestingly, if the gold surface of a surface plasmon resonance imaging (SPRi) prism was used for the formation of SAMs through thiol or disulfide bonds, a label-free, synchronous, parallel, and real-time observation of binding events on the CoCRRs array could also be performed (Figure 1). To investigate this hypothesis, we designed a model array inspired by the way cell-surface heparan sulfates (HS) recognize HS binding proteins (HSbps), an important group of extracellular mediators involved in many physiological and pathological processes. HS are negatively charged polysaccharides displaying a high degree of molecular diversity, which arises from the regulated generation of various epimerization and sulfation patterns during biosynthesis. According to cell type and activation state, HS chains with different negatively charged topologies are expressed, presumably to promote selective interactions with HSbps. [*] Dr. Y. Hou, M. Genua, Dr. D. Tada Batista, Dr. R. Calemczuk, Dr. A. Buhot, Dr. T. Livache SPrAM, UMR 5819 (CEA-CNRS-UJF-Grenoble 1), INAC/CEA-Grenoble 38054 Grenoble cedex 9 (France) E-mail: yanxia.hou-broutin@cea.fr thierry.livache@cea.fr

Artificial Ion Channel Biosensor in Human Immunodeficiency Virus gp41 Drug Sensing

An ion channel biosensor is described for label-free detection of inhibitors which bind to the coiled coil domain of human immunodeficiency virus type 1 gp41. gp41 is the viral transmembrane glycoprotein responsible for fusion between HIV-1 and host cells. The N-terminal coiled coil domain binds three antiparallel C-heptad repeat peptides in the six helix bundle structure of trimeric gp41 that forms during fusion. Compounds able to prevent six-helix bundle formation by binding to the gp41 coiled coil could inhibit fusion and have important therapeutic potential. We have immobilized on gold a positively charged metallopeptide mimic of a section of the gp41 coiled coil containing a hydrophobic pocket suitable for small molecule binding. We demonstrate that the resulting sensor is able to transmit a current in the presence of negatively charged redox marker ions, therefore acting as an artificial ion channel. The electrochemical signal, measured by cyclic voltammetry, was modulated by specific analyte binding to the coiled coil. Nanomolar quantities of peptides and small molecules that bind in the hydrophobic pocket could be selectively detected, providing a method for label-free detection of binding to gp41.

A Versatile Electronic Tongue Based on Surface Plasmon Resonance Imaging and Cross-Reactive Sensor Arrays—A Mini-Review

Nowadays, there is a strong demand for the development of new analytical devices with novel performances to improve the quality of our daily lives. In this context, multisensor systems such as electronic tongues (eTs) have emerged as promising alternatives. Recently, we have developed a new versatile eT system by coupling surface plasmon resonance imaging (SPRi) with cross-reactive sensor arrays. In order to largely simplify the preparation of sensing materials with a great diversity, an innovative combinatorial approach was proposed by combining and mixing a small number of easily accessible molecules displaying different physicochemical properties. The obtained eT was able to generate 2D continuous evolution profile (CEP) and 3D continuous evolution landscape (CEL), which is also called 3D image, with valuable kinetic information, for the discrimination and classification of samples. Here, diverse applications of such a versatile eT have been summarized. It is not only effective for pure protein analysis, capable of differentiating protein isoforms such as chemokines CXCL12α and CXCL12γ, but can also be generalized for the analysis of complex mixtures, such as milk samples, with promising potential for monitoring the deterioration of milk.

SPR imaging based electronic tongue via landscape images for complex mixture analysis.

Electronic noses/tongues (eN/eT) have emerged as promising alternatives for analysis of complex mixtures in the domain of food and beverage quality control. We have recently developed an electronic tongue by combining surface plasmon resonance imaging (SPRi) with an array of non-specific and cross-reactive receptors prepared by simply mixing two small molecules in varying and controlled proportions and allowing the mixtures to self-assemble on the SPRi prism surface. The obtained eT generated novel and unique 2D continuous evolution profiles (CEPs) and 3D continuous evolution landscapes (CELs) based on which the differentiation of complex mixtures such as red wine, beer and milk were successful. The preliminary experiments performed for monitoring the deterioration of UHT milk demonstrated its potential for quality control applications. Furthermore, the eT exhibited good repeatability and stability, capable of operating after a minimum storage period of 5 months.

Fabrication of nanotweezers and their remote actuation by magnetic fields

A new kind of nanodevice that acts like tweezers through remote actuation by an external magnetic field is designed. Such device is meant to mechanically grab micrometric objects. The nanotweezers are built by using a top-down approach and are made of two parallelepipedic microelements, at least one of them being magnetic, bound by a flexible nanohinge. The presence of an external magnetic field induces a torque on the magnetic elements that competes with the elastic torque provided by the nanohinge. A model is established in order to evaluate the values of the balanced torques as a function of the tweezers opening angles. The results of the calculations are confronted to the expected values and validate the overall working principle of the magnetic nanotweezers.

Electronic tongue generating continuous recognition patterns for protein analysis.

In current protocol, a combinatorial approach has been developed to simplify the design and production of sensing materials for the construction of electronic tongues (eT) for protein analysis. By mixing a small number of simple and easily accessible molecules with different physicochemical properties, used as building blocks (BBs), in varying and controlled proportions and allowing the mixtures to self-assemble on the gold surface of a prism, an array of combinatorial surfaces featuring appropriate properties for protein sensing was created. In this way, a great number of cross-reactive receptors can be rapidly and efficiently obtained. By combining such an array of combinatorial cross-reactive receptors (CoCRRs) with an optical detection system such as surface plasmon resonance imaging (SPRi), the obtained eT can monitor the binding events in real-time and generate continuous recognition patterns including 2D continuous evolution profile (CEP) and 3D continuous evolution landscape (CEL) for samples in liquid. Such an eT system is efficient for discrimination of common purified proteins.

Mode of PEG Coverage on Carbon Nanotubes Affects Binding of Innate Immune Protein C1q

Surface modification of nanoparticles with poly(ethylene glycol) (PEG) is used in biomedicine to increase the circulation time of the particles after intravenous injection. Here, we study the interaction of PEG-covered carbon nanotubes (CNTs) with the serum complement protein C1q. Besides being the target-recognizing unit of the initiating complex for the classical pathway of complement in our innate immune system, C1q is involved in a range of important physiological processes. We modified the surface of multiwalled CNTs with covalently grafted PEG and physically adsorbed PEG. Transmission electron microscopy revealed the interaction of these PEG-coated CNTs with C1q. We found abundant C1q coverage on the PEG-grafted CNTs but not on the CNTs with adsorbed PEG. We tested the ability of these CNTs to activate the complement system using in vitro complement activation assays. None of the CNTs studied activated the C1q-dependent classical complement pathway. These findings are pertinent to the safe design and novel biomedical applications of PEGylated CNTs.