第一回　光・プラズマプロセスのバイオ応用ワークショップ

The 1st Workshop on Biological Applications of Plasma/Photon Processing


Abstracts



PLASMA PROCESSED SURFACES FOR SELECTED CELL RESPONSES
P. Favia, E. Sardella, R. Gristina, M. Nardulli, L.C. Lopez, E.
Dilonardo, B. Pistillo, R. d’Agostino
Department of Chemistry, University of Bari, Italy
Institute of Inorganic Methods and Plasmas, CNR, Bari, Italy
Plasma deposited coatings and plasma treated surfaces can induce pre-determined properties at the surface of polymers and other materials, that are desirable for in vitro, ex vitro and in vivo biomedical applications such as biomaterials, implantable prostheses, biosensors, tissue engineering and biomedical devices. This talk will describe plasma functionalization processes that have been optimized for obtaining bacterial-resistant coatings, functional surfaces for biomolecule immobilization and surfaces with selected acid/basic character, that have been utilized to induce selected responses by cells and bacteria.


Developments of Ionchannel Biosensor and Plasma Process
「イオンチャンネルバイオセンサーとプラズマプロセスの開発」
Tsuneo Urisu
宇理須恒雄
Institute for Molecular Science/SOKENDAI
分子科学研究所/総合研究大学院大学
The membrane protein such as ion channel and GPCR plays an important role as a receptor of the information in vivo transmission. It accounts for 50% or more of all drug development targets, because it takes part in almost all diseases called an intractable disease. However, since there is no excellent high-through-put screening biosensor for the membrane protein, this makes the research and developments of new drugs difficult. In this work, we have developed a planer type ion channel biosensor using a Si substrate with micro pore、where TRPV1–channel-expressed HEK293 cell is positioned. Ligand gated ion channel currents have been successfully observed using capsaicin as a ligand molecue. Because the micro-fabrication of the Si substrate is the most important process, the related plasma process is described.
　膜タンパクは、生体内の情報伝達の受信素子として重要な役割を果たしており、殆どの、難病と言われる疾患に関与しているため、　全創薬ターゲットの50％以上を占める。　しかし、高性能な、ハイスループットスクリーニング用のバイオセンサーが無いため、創薬研究開発を困難にしている大きな原因の一つである。　本研究では、膜タンパクであるイオンチャンネルのバイオセンサーとして、HEK293 細胞にTRPV1チャンネルを発現し、これを微細貫通孔を有するSi基板に搭載したプレーナー型イオンチャンネルバイオセンサーを開発し、カプサイシンのリガンド刺激電流の検出に成功した。　Si基板の微細加工が最も重要な工程として、関連のプラズマプロセスについて述べる。

Nano-carbon Films Formed with Electron Cyclotron Resonance (ECR) Sputtering and Their Application for Bioanalysis and Biosensors
Osamu Niwa

National Institute of Advanced Industrial Science and Technology (AIST)
Division of Biological Resources and Functions
　The carbon thin films deposited by electron cyclotron (ECR) sputtering method have unique structure and properties. The films consist of sp3 and sp2 mixed bonds and have equivalent mean scratch hardness to the diamond. The structure of the film from TEM image is different from usual graphite carbon. The content of sp3 bonds increases with increasing acceleration voltage during sputtering. As an electrode for electroanalysis, the film shows wide potential window and low capacitive current and therefore applied for measuring various biomolecules with high oxidation potentials including histamine and NADPH and nucleotides. We prepared biosensor for detecting -aminobutyric acid (GABA) which is a well know inhibitory neutrotransmitter by modifying ECR sputtered carbon film with enzyme layer. The sensor shows a low detection limit which is more than 1 order of magnitude of lower than that of biosensor based on the conventional glassy carbon (GC) electrode. With ECR sputtered carbon film electrodes, we also succeeded in determining the oxidation peak potentials for 4 kind of nucleotides (GMP:1.23V, AMP:1.57V, TMP:1.78V, CMP:1.99 V). In addition, the content of bases in oligonucleotide can be roughly measured by differential pulse voltammetry (NPV) using the ECR sputtered carbon film electrodes since the decrease in the oxidation currents caused by the adsorption of oligonucleotide can be suppressed due to weak interaction between the analytes and electrode surface.

Near field Raman spectroscopy for molecular nano-imaging and nano-
analysis
Yasushi Inouye
Graduate School of Frontier Biosciences, Osaka University
Plasomon polaritons, collective oscillation of electrons coupled with photons, are generated in metallic nanostructures when light is incident on the nanostructures. Then, photons are confined and enhanced in the vicinity of the nanostructures. We proposed a metallic needle having a nano-apex as nano-light source for microscopy having super resolving capability, which is called as near field microscopy. In this presentation, we will show Raman nanospectroscopy by using this microscopic technique for molecular nano-imaging and nano-analysis. Furthermore we will discuss nonlinear effects generated in near field Raman spectroscopy which provides us spatial resolution exceeding 10 nm.

Molecular Dynamics Simulations of Cell Membrane Nanopores with DNA
Motohiko Tanaka, and Yitzak Rabin
Electrostatic effects sometimes play important roles in structure formation of biological systems, most notable of which is seen in a tight structure of double helix DNA. Here we talk about our molecular dynamics study of membrane nanopores with or without a single-stranded(ss) DNA inside [1,2]. The cell membrane is characterized by very low dielectric constant (2) compared to that of surrounding water (80), which leads to dramatic phenomena due to electrostatic effects, including exclusion of isolated charged ions and/or paring of salt ions in nanopores. A ss-DNA takes a stretched configuration in the nanopore due to electrostatic repulsion from the pore wall.
References
[1] Y.Rabin and M.Tanaka, Phys.Rev.Lett., 94, 148103 (2005).
[2] M.Tanaka and Y.Rabin, Flow Dynamics, pp.212-216 (AIP CP832, 2006).
Also, refer to http://dphysique.nifs.ac.jp/

Construction of Patterned Cell Spheroid Array for High Performance Cell Based Biosensor
Yukio Nagasaki
Tsukuba Research Center for Interdisciplinary Materials Science (TIMS), and Center for Tsukuba Advanced Research Alliance (TARA),
University of Tsukuba, Ten-noudai 1-1-1, Tsukuba, Ibaraki 305-8573, Japan
In order to create high performance biofunctionality devices such as protein, DNA and cell array chips, it is important to control an adsorption of biomolecules and cells on substrate surface. The objective of this work was to construct patterned cell culture system on precisely designed micropattened biointerface, which was constructed by our original surface modification method using PEG derivatives. The constructed patterned cell spheroid array, using primary hepatocyte celle is promising as new cell based biosensor.

Material processing by atmospheric plasma jets and plasmas in contact with liquids
Katsuhisa Kitano and Satoshi Hamaguchi
Center for Atomic and Molecular Technologies, Graduate School of Engineering, Osaka University
Many types of atmospheric pressure plasma jets (APPJs) have been developed. At the atmospheric pressure, due to frequent collisions between high-energy plasma particles and neutral gas, discharges can be easily transformed to thermal plasmas (e.g., arc plasmas). Nevertheless nonthermal (nonequilibrium) plasmas are also achieved by several techniques. From a viewpoint of plasma applications to materials, such plasmas offer attractive opportunities since nonthermal plasmas can supply chemically active species without heating and/or damaging the materials to be processed. Characteristics of nonthermal APPJs generated in and/or from a dielectric tube which is wrapped around by two tubular electrodes connected to a low frequency (~10kHz) high voltage power supply [M. Teschke, et al., IEEE. trans. Plasma Sci., 33, 310, (2005)] and other related systems are discussed. The aim of this study is to understand the mechanism of such nonequilibrium APPJ and also to extend the concept for the development of new plasma generation systems at atmospheric pressure.