1. Construction of On-chip Biosensing Devices
Biosensing systems, such as enzyme, immuno sensors and DNA micro arrays, are widely used in the field of medical care and medicine manufacturing. In particular, recent progress in genome engineering requires high performance integrated micro-multi biosensing system, which can be utilized for recognition of individual biomolecule and analysis of bioreaction at single molecular level.
In order to realize highly sensitive biosensing system, precise fabrication of the electrode parts for molecular recognition is a significant issue. For this, developments of new detection devices with high sensitivity are strongly demanded. Especially, it is desired that the electrode surface has the supermolecular structure that minics cell systems. In order to fabricate such an electrode, application of the template for the ordered-arrangement of the molecules is effective. Organic monolayers have self-assemble ability onto the surfaces; the monolayer modified electrode is suitable as the templates for orderly immobilization of biomolecules. On the other hand, it is preferable that the detection system can detect the signal immediately and high sensitively. A field-effect transistor type electrode can directly detect surface reactions as an electric signal, with capability for the on-chip integration.
In the present work, to construct of highly sensitive on-chip integrated micro-biosensing deveces, we investigate the formation of functionalized organic monolayer modified electrode on silicon wafer surfaces and the development of the new detection system utilizing semiconductor device such as a field effect transistor.
Fig. 1-1 "Schematic design of on-chip semiconductor/organic monolayer transistor base biosensing device.
Figure 1-1 shows basic design of the on-chip integrated biosensing devices. In order to fabricate these devices, the fabrication of the transistor that has chemical resistance and the position selective formation of the organic monolayer having different functional group on each gate electrode are required. At the sensing electrode, amino functionalized monolayer is suitable for immobilization of DNA, enzyme, etc. For the reference electrode, an alkyl, or perfuloro-alkyl functionalized monolayer is effective. Hence, the formation of various functionalized monolayer and its patterning are one of the key processes for the fabricating on chip biosensing devices.
Fig. 1-2 Schematic design of organic monolayer template.
Currently, we are investigating the formation of "organic monolayer templates" which are position selectively formed with the monolayers having various functional groups (Figure 1-2) and the immobilization of biomolecules onto the templates. From the result, it is indicated that DNAs are immobilized selectively only to the active sites (amino sites) on the templates. Moreover, we attempt the design and development of fabrication process for the feild effect transistor with high chemical durability.
Related Paper(s)
¥ Daisuke Niwa, Kaoru Omichi, norikazu Motohashi, Takayuki Homma, Tetsuya Osaka, "Organosilane self-assembled monolayer-modified field effect transistors for on-chip ion and biomolecule sensing", Sens. Actuator B-Chem., 108, 721-726 (2005).
¥ D. Niwa, K. Omichi, N. Motohashi, T. Homma, T. Osaka, "Formation of Micro and Nanoscale Patterns of Monolayer Templates for Position Selective Immobilization of Oligonucleotide Using Ultraviolet and Electron Beam Lithography", Chem. Lett., 33, 2, 176-177 (2004).
¥ D. Niwa, Y. Yamada, T. Homma, T. Osaka, "Formation of Molecular Templates for Fabricating On-Chip Biosensing Devices", J. Phys. Chem. B, 108, 10, 3240-3245 (2004).
2. Fabrication and Characterization of Chiral Surfaces for Chiral Sensors
Many biologically active substances occur in the form of mirror-image isomers called enantiomers. Nature mainly uses one of the two enantiomers. All living material have amino acids, and therefore peptides, enzymes and other proteins, only of one of the enantiomeric forms. An understanding of chirality induction and discrimination is an important issue in the fields of chemistry, physics, biology, and pharmacy. In particular, an induction of chirality to a solid surface is expected to be very useful for the application to a sensor for chiral compounds and an enantioselective electrode or heterogeneous catalyst.
As the origin of enantioselectivity, the chirality of molecular arrangement should be considered as well as that of the molecule itself, especially in the case where the surface consists not of a simple random assembly of chiral compounds but of a two-dimensionally ordered arrangement of molecules. Aiming at an application to sensing devices for chiral discrimination, we are investigating the enantioselectivity of the solid surface modified with chiral molecules.
A self-assembled monolayer (SAM) of 1,1'-binaphthalene-2,2'-dithiol (BNSH) was reported to consist of a well-ordered two-dimensional arrangement on a gold(111) single-crystal surface. We investigate the adsorption behaviors of chiral analytes on BNSH SAM by using a quartz crystal microbalance (QCM) technique. The enantioselective adsorption of chiral amino acid, phenylalanine, onto BNSH SAMs was detected and the two-dimensional chiral arrangement was suggested to play an important role in the enantioselectivity of adsorption of phenylalanine. Applications of the present system to other analyte molecules are under investigation.
Fig. 2-1 Characterization of enantioselectivity of the surface by QCM.
Development of sensing devices for chiral compounds, such as an optical-purity sensor, is our final goal. We are aiming at the fabrication of highly-enantioselective system in contrast to most of the chiral sensors reported so far showing responses not only to the target enantiomer but also the other one.
We have found that the crystal growth of chiral amino acid on a solid surface modified with the SAM, to which enantiomeric molecules are covalently attached, occurs enantioselectively, as evidenced by the X-ray diffraction method. Based on the quantitative detection of this phenomenon by QCM, a practical approach to the development of novel chiral sensors is now in progress.
Fig. 2-2 Chiral sensing based on enantioselective crystal growth.
Related Paper(s)
¥ N. Banno, T. Nakanishi, M. Matsunaga, T. Asahi, T. Osaka, "Enantioselective Crystal Growth of Leucine on a Self-Assembled Monolayer with Covalently Attached Leucine Molecules", J. Am. Chem. Soc., 126, 2, 428-429 (2004).
¥ T. Nakanishi, N. Yamakawa, T. Asahi, N. Shibata, B. Ohtani, T. Osaka, "Chiral Discrimination Between Thalidomide Enantiomers Using a Solid Surface", Chirality, 16, S1, S36-S39 (2004).
2. Fabrication and Characterization of Chiral Surfaces for Chiral Sensors
Magnetic nanoparticles have been attracting much interest as a labeling material in the fields of advanced biological and medical applications such as drug delivery, magnetic resonance imaging, and array-based assaying. In these cases, nanoparticles of iron oxides such as magnetite (Fe3 O4 ) and maghemite (ƒÁ-Fe2 O3 ) need to be modified with organic molecules for immobilization of biological materials.
Controlling the affinity to biological materials and the dispersibility to solvents becomes very important as well as a control of the particle size and the magnetic property of iron oxide nanoparticles. These features are influenced by preparation techniques because of their dependence on the particle size, size distribution, and surface properties. The pyrolysis of an organometallic compound and the coprecipitaion by alkalization of a solution of metal salt are commonly used as a chemical preparation technique for iron oxide nanoparticles. We carried out a unique preparation of iron oxide nanoparticles in reverse micelles. Nanoparticles of ƒÁ-Fe2 O3 were synthesized via reduction of iron ionic species by reducing agents and subsequent air oxidation in the nano-sized water droplet in reverse micelles. The obtained nanoparticles of ƒÁ-Fe2 O3 with 4 nm in diameter exhibited a relatively high crystallinity, a narrow size distribution, and a superparamagnetic behavior. Furthermore, the surface modification of nanoparticles with the organic molecule containing an amino group is in progress for the immobilization of biological materials to nanoparticles.
In addition, we also investigate the application of magnetic nanoparticles to the biosensing. The detection of the area-selected immobilization of magnetic nanoparticles onto the substrate patterned with self-assembled monolayers based on the specific interaction between biotin and streptavidin was demonstrated by magnetic force microscopy (MFM). The application of the magnetic detection method to the antigen-antibody system is under investigation for the use in the field of medical care.
Related Paper(s)
¥ T. Nakanishi, H. Iida, T. Osaka, "Preparation of Iron Oxide Nanoparticles via Successive Reduction-Oxidation in Reverse Micelles", Chem. Lett., 32, 12, 1166-1167 (2003).
¥ H. Iida, T. Nakanishi, T. Osaka, "Surface Modification of ƒÁ-Fe2O3 Nanoparticles with Aminopropylsilyl Groups and Interparticle Linkage with ƒ¿,ƒÖ-Dicarboxylic Acids", Electrochim. Acta, 51, 855-859 (2005).
¥ A. Arakaki, S. Hideshima, T. Nakagawa, D. Niwa, T. Tanaka, T. Matsunaga, T. Osaka, "Detection of Biomolecular Interaction Between Biotin and Streptavidin on a Self-Assembled Monolayer Using Magnetic Nanoparticles", Biotechnol. Bioeng., 88, 4, 543-546 (2004).
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