Toshikazu Kawaguchi

Fabrication of a novel immunosensor using functionalized self-assembled monolayer for trace level detection of TNT by surface plasmon resonance

We have developed a new immunosensor based on self-assembly chemistry for highly sensitive and label-free detection of 2,4,6-trinitrotoluene (TNT) using surface plasmon resonance (SPR). A monolayer of amine terminated poly(ethylene glycol) hydrazinehydrochloride (PEG-NH(2)) thiolate was constructed on an activated gold surface and immobilized with trinitrophenyl-beta-alanine (TNPh-beta-alanine) by amide coupling method. The binding interaction of a monoclonal anti-TNT Ab (M-TNT Ab) with TNPh-beta-alanine immobilized thiolate monolayer surface was monitored and evaluated for detection of TNT based on the principle of indirect competitive immunoreaction. Here, the competition between the self-assembled TNT derivative and the TNT in solution for binding with antibody yields in the response signal that is inversely proportional to the concentration of TNT in the linear detection range. With the present immunoassay format, TNT could be detected in the concentration range from 0.008ng/ml (8ppt) to 30ng/ml (30ppb). The response time for an immunoreaction was 2min and one immunocycle could be done with in 4min including surface regeneration. Bound antibodies could be easily eluted from the self-assembled immunosurface at high recoveries (more than 100 cycles) using pepsin solution without any damage to the TNT derivatives immobilized on the surface. The compact self-assembled monolayer was highly stable and prevented the non-specific adsorption of proteins on the surface favoring error free measurement.

Preparation of binary metal electrocatalysts by self-assembly of precursor ionic species on gold and reduction of nitrate ions

Pt/Sn binary metal electrodes were prepared by the successive adsorption of Sn2+ and PtCl62− ions onto a gold substrate followed by the reduction of the adsorbed PtCl62−. The atomic ratios of Sn and Pt to the surface Au were determined to be approximately 1 and 0.3, respectively, using a quartz crystal microbalance (QCM), X-ray photoelectron spectroscopy (XPS) and cyclic voltammetry. Therefore, binary metal layers at a monolayer level were formed. These Pt/Sn binary metal electrodes exhibited a very high electrocatalytic activity for the reduction of nitrate ions with the predominant formation of ammonium ions.

Development of a sensitive surface plasmon resonance immunosensor for detection of 2,4-dinitrotoluene with a novel oligo (ethylene glycol)-based sensor surface.

A surface plasmon resonance (SPR) immunosensor for detection of 2,4-dinitrotoluene (2,4-DNT), which is a signature compound of 2,4,6-trinitrotoluene-related explosives, was developed by using a novel oligo (ethylene glycol) (OEG)-based sensor surface. A rabbit polyclonal antibody against 2,4-DNT (anti-DNPh-KLH-400 antibody) was prepared, and the avidity for 2,4-DNT and recognition capability were investigated by indirect competitive ELISA. The sensor surface was fabricated by immobilizing a 2,4-DNT analog onto an OEG-based self-assembled monolayer formed on a gold surface via an OEG linker. The fabricated surface was characterized by Fourier-transform infrared-refractive absorption spectrometry (FTIR-RAS). The immunosensing of 2,4-DNT is based on the indirect competitive principle, in which the immunoreaction between the anti-DNPh-KLH-400 antibody and 2,4-DNT on the sensor surface was inhibited in the presence of free 2,4-DNT in solution. The limit of detection for the immunosensor, calculated as three times the standard deviation of a blank value, was 20 pg mL(-1), and the linear dynamic range was found to be between 1 and 100 ng mL(-1). Additionally, the fabricated OEG-based surface effectively prevented non-specific adsorption of proteins, and the specific response to anti-DNPh-KLH-400 antibody was maintained for more than 30 measurement cycles.

Underpotential deposition of zinc ions on polycrystalline platinum: FTIR and EQCM study

Abstract Underpotential deposition(UPD) of Zn 2+ ions on polycrystalline platinum in KH 2 PO 4 solution (pH 4.6) was studied by Fourier transform infrared reflection spectroscopy (FTIR) and electrochemical quartz crystal microbalance (EQCM). By the addition of Zn 2+ ions in KH 2 PO 4 solution, the intensities of ir bands of around 990, 1044 and 1090 cm −1 are increased with a negative increase of potentials at E ⩽ 0.7V ( rhe ) where Zn UPD takes place, while by EQCM the weight increase of Pt mounted quartz was observed in the same potential region. Coverage of UPD Zn was 0.75 at 0.05 V as two-electron transfer UPD process, being estimated from cyclic voltammogram, and mps (mass number per platinum sites) was 78 by EQCM; from both values the coverage of phosphate was evaluated as 0.27. These results are discussed that the phosphate species adsorbs over UPD Zn on Pt as a form of HPO 4 , where one of three Pue5f8Os seems to be bound to UPD Zn on Pt.

Highly selective and sensitive detection of β-agonists using a surface plasmon resonance sensor based on an alkanethiol monolayer functionalized on a Au surface.

Immunosensor surfaces for surface plasmon resonance (SPR) have been constructed using a functionalized succinimidyl propanethiol monolayer as a linker to immobilize β-agonist protein conjugates on a Au surface. Because β-agonist is a small molecule, an indirect competitive inhibition immunoassay was used for detection. The lowest detection limits for ractopamine and salbutamol were 10 ppt (10 pg mL(-1)) and 5 ppt (5 pg mL(-1)), respectively. The fabricated immunosensor surface can be used again for detection after regeneration in 0.1 M sodium hydroxide. It was found that the same sensor surface could be reused for performing over 100 rapid immunoreactions. Moreover, one immunosensing-regeneration cycle requires only 600 s. The fabricated immunosensor surfaces were characterized using SPR and scanning tunneling microscopy observation. In the kinetic study of the indirect competitive immunosensing inhibition, the affinity constant (K1) of salbutamol antibody was smaller than the K1 of ractopamine antibody. Compared to a previous study of clenbuterol detection, it was concluded that the high K1 was coupled with low sensitivity. In the selectivity study, both immunosensor surfaces provided >90% of confidence level for the specific detection of β-agonist compounds. The fabrication of highly selective and sensitive sensor surfaces for detecting β-agonist compounds was confirmed.

Improvement in Propene Sensing Characteristics by the Use of Additives to In2O3 Sensing Electrode of Mixed-Potential-Type Zirconia Sensor

In an effort to improve propene (C 3 H 6 ) selectivity and long-term stability of the mixed-potential-type yttria-stabilized zirconia (YSZ)-based tubular sensor attached with In 2 O 3 sensing electrode (SE), each of the nanosized noble metals (Pt, Ru, Pd, Rh, or Ir) and YSZ powder was added to In 2 O 3 SE. It was found that the addition of nanosized Pt (0.3 wt %) gave considerable improvement in C 3 H 6 selectivity, and the addition of YSZ powder (10 wt %) exhibited great improvement in long-term stability of the sensor. Thus, the sensor attached with composite SE consisting of In 2 O 3 , 0.3 wt % Pt, and 10 wt % YSZ was fabricated and its sensing characteristics to C 3 H 6 were examined. As a result, the present sensor exhibited high selectivity and excellent long-term stability to C 3 H 6 at 450°C under the wet condition for the examined period of about 1 month. Based on the results obtained from the measurement of catalytic activity for CO oxidation to CO 2 , the observation of morphology, and the analysis of composition of SEs, it was speculated that the added Pt nanoparticles could promote the oxidation of CO and H 2 and then improve the C 3 H 6 selectivity, while the added YSZ powder could stabilize the morphology of the SE/YSZ interface and give better long-term stability.

Redox and mass transport characteristics of domain-free mixed ferrocenyloctanethiol/alkanethiol monolayers on gold

Mixed monolayers of ferrocenyloctanethiol (FcC8SH) and alkanethiols of various chain lengths were constructed after predetermining the surface composition by the initial coverage of Pb underpotentially deposited on a bare gold electrode during the first step of the preparation. Single waves observed in the reductive desorption voltammograms show no formation of single-component domains. These mixed monolayers showed the following characteristics. First, a considerable amount of the ferrocene moieties were not electroactive. Second, the mass change during the redox reaction of the ferrocene was greater than that expected from the simple association of the ferrocenium cation with the electrolyte anion, and that for the mixed monolayers prepared from mixed solutions of the precursor thiols. This large mass change was interpreted by the incorporation of the anion and water upon oxidation into the pocket surrounded by alkanethiol and FcC8SH at the bottom. These results are consistent with those expected for the FcC8SH molecules in a domain-free environment, or surrounded by more alkanethiol molecules than FcC8SH molecules.

Construction of mixed mercaptopropionic acid/alkanethiol monolayers on polycrystalline gold electrodes using underpotentially deposited lead as the control element

The method we proposed in our previous paper (J. Am. Chem. Soc. 124 (2002) 654) for the construction of mixed monolayers on Au(111) was applied to polycrystalline Au. X-ray photoelectron spectroscopy, Fourier transform infrared spectroscopy and reductive desorption voltammetry demonstrate that the surface coverage of mercaptopropionic acid (MPA) in mixed monolayers with alkanethiol was controlled by the initial coverage of Pb underpotentially deposited during the first step of the preparation. In addition, MPA and alkanethiol did not form single-component domains, the sizes of which are large enough to create two distinct sets of reductive desorption waves. In contrast, single-component domains were formed when mixed monolayers were formed from mixed solutions.

Integrated microfluidic device for the separation and electrochemical detection of catechol estrogen-derived DNA adducts

Catechol estrogen-derived DNA adducts are formed as a result of the reaction of catechol estrogen metabolites (e.g., catechol estrogen quinones) with DNA to form depurinating adducts. Developing a new methodology for the detection of various DNA adducts is essential for medical diagnostics, and to this end, we demonstrate the applicability of on-chip capillary electrophoresis with an integrated electrochemical system for the separation and amperometric detection of various catechol estrogen-derived DNA adducts. A hybrid PDMS/glass microchip with in-channel amperometric detection interfaced with in situ palladium decoupler is utilized and presented. The influence of buffer additives along with the effect of the separation voltage on the resolving power of the microchip is discussed. Calibration plots were constructed in the range 0.4–10xa0μM with r2u2009≥u20090.999, and detection limits in the attomole range are reported. These results suggest that on-chip analysis is applicable for analyzing various DNA adducts as potential biomarkers for future medical diagnostics.

Unusual catalytic effect of the two-dimensional molecular space with regular triphenylphosphine groups.

A novel organic-inorganic hybrid 2D molecular space with regular triphenylphosphine groups (triphenylphosphineamidephenylsilica, PPh(3)APhS) was successfully synthesized through grafting triphenylphosphine groups in the 2D structure of layered aminophenylsilica dodecyl sulfate (APhTMS-DS), which was developed in our previous research, with regular ammonium groups. The 2D structures were kept after the grafting reaction of triphenylphosphine groups in PPh(3)APhS. The catalytic potentials of 2D molecular space with regular triphenylphosphine groups were investigated. An unusual catalytic effect was found in a carbon-phosphorus ylide reaction. The PPh(3)-catalyzed reaction of modified allylic compounds, including bromides and chlorides with tropone yielded a [3 + 6] annulation product. However, an unusual [8 + 3] cycloadduct was obtained in the reaction of modified allylic compounds, including bromides and chlorides with tropone catalyzed by PPh(3)APhS. Otherwise, the stable catalytic intermediate was successfully separated, and the reaction activity of the catalytic intermediate was confirmed in the reaction of modified allylic compounds with tropone catalyzed by PPh(3)APhS. This research is the first successful example of directly influencing catalytic reaction processes and product structures by utilizing the chemical and geometrical limits of 2D molecular spaces with regular catalyst molecules and affords a novel method for controlling catalytic reaction processes and catalyst design.