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國科會補助專題研究計畫出席國際學術會議心得報告
日期：101 年 07 月 31 日
一、參加會議經過
IEEE NEMS Conference 是一有關微工程技術交流之國際研討會，其領域包括生醫感測器、半導體 製程及最新的生物領域相關技術，每年度於世界各地舉辦一次，與會人士皆為世界各地之頂尖學者及 研究人員；今年會議日期為 2012/3/5 至 2012/3/8 共四天，主辦單位為京都大學，會議則於日本的京都 舉行，京都有悠久歷史為日本獲選世界遺產的城市之一，其建築群主要效仿隋唐長安城與洛陽城設計 古稱平安京，於桓武天皇時期完成(西元 794 年)定為國都直到明治天皇(西元 1868 年)遷都東京為止，
為日本歷經千年的政治及文化的中心。
第一天主要於京都大學內辦理報到相關事宜，參加者領取名牌及會議相關資料。第二天至第四天 則為正式議程，包含每日九點開始的例行的研討會議，每日會議歷程分上午與下午共有四個場次，此 外除每日例行研討會議也於午後時段舉行海報報告，讓與會者皆能找到感興趣之研究主題，此次會議 共計有 230 場的口頭報告與 104 場的海報報告。
本實驗室在此會議共發表兩篇論文，其中”Development of Microbead-based Affinity Biosensor by Insulator-Based Dielectrophoresis”是安排為口頭報告論文，於會議的第三天下午，由本實驗室研究生卓
計畫編號 NSC 100－2221－E－009－034－
計畫名稱 以高分子作為感應偶合電漿反應離子蝕刻側壁保護以製作單
晶矽懸浮微結構之快速製程平台研發 出國人員
姓名 徐文祥、卓琮閔 服務機構
及職稱 交通大學機械系教授、研究生
會議時間
2012 年 03 月 05 日 至
2012 年 03 月 08 日
會議地點 日本，京都
會議名稱
(中文)奈米/微米工程與分子系統國際研討會
(英文)2012 7th IEEE International Conference on Nano/Micro Engineered and Molecular Systems
發表題目
共兩篇
Development of Microbead-based Affinity Biosensor by Insulator-Based Dielectrophoresis
Fabrication of Deep Lateral Single-Crystal-Silicon Blaze Micro-grating by
Inductively-Coupled-Plasma Reactive Ion Etch
並向負責報告的研究生討論研究相關內容，而能透過國際研討會與相關研究領域的學者交談，也讓學 生獲益良多。另一篇”Fabrication of Deep Lateral Single-Crystal-Silicon Blaze Micro-grating by
Inductively-Coupled-Plasma Reactive Ion Etch”，則是安排為壁報論文，也在壁報展示時間同與會學者交 流，獲得寶貴經驗。
而於第三天的晚上所舉辦的餐會，除了可讓與會者放鬆心情外，並可得到更進一步交流的機會，
本實驗室也於晚上餐會中，與結識的數位學者交流有關研究上的各種心得；另外主辦單位也在餐會中，
向與會者宣傳下一屆研討會的資訊，包括會議的日期、地點以及當地特色，期許眾人能於下屆研討會 中，交流更多的研究成果。
二、與會心得
此次研討會中，本實驗室研究生以英文向國際學者報告研究成果，報告情形如圖一所示。另外雖 然受限於時間，無法對研究內容作更進一步的詳細介紹，但對相關領域感興趣之學者，也於報告時間 之後做進一步的交流討論，如圖二所示。
分享研究成果的過程實在令人感到相當興奮，是一種自我實現的成就感，而透過學術交流世界接 軌的過程更是難能可貴。在研討會中各個會議報告都是其精心研究的成果，而利用此次國際交流的機 會，吸收這次研討會所提出的嶄新想法，勢必會對學生日後的研究產生很大的影響力。
此外於一些生醫感測器、生醫化學、半導體製程技術等領域上的收穫也頗豐碩，直到最後一天時 仍有意猶未盡之感，希望時間能持需停留於此刻。此次參加研討會外參訪的當地人文風情、增進了學 術與人文的素養，然而最有趣也最有意義的是進行了許多國民外交，無論是一同前往的教授們、國外 學者、對岸朋友或是台灣學生。
圖一、本實驗室研究生於會場內進行報告之照片
圖二、報告結束時與學者討論之照片
在會場也遇到多位台灣教授，一同參與此研討會，如圖三所示，左邊第一位是本人(徐文祥)，右邊 第二位是交大電子系鄭裕庭教授，透過討論聽講心得，得到更豐富的經驗分享。
圖三、參加會議之學者合照
三、建議
參加研討會不僅僅是學習，更可以開拓視野深度，探訪當地的風土及其民族精神，這完全取決於 心態，唯有將自我放空與縮小，才能吸收更多養分回國深究。在國內告訴研究生該做哪些準備，學生 常不能體會原因，透過親身經歷，學生更能體會為何要在出國前，事先將所有口頭報告與海報報告的 標題進行了解，挑出有興趣的題目後，到了會場便可快速地進入狀況，迅速地拜訪每個學者，這跟平 常在實驗室閱讀文獻有很大的不同，透過當面的問與答可以加速資訊的吸收之外，更可獲取一些文獻 中沒有包含到的資訊，對於研究工作相當有幫助；此外也可同時留意國內一同與會的學者，即使研究 領域不同，但也可以於會場中結識而替未來的合作留下一個種子。
四、攜回資料名稱及內容
研討會紀念提袋，內含：
(1)議程簡章
(2)會議手冊(內含報告主題與摘要) (3)光碟(內含論文電子檔)
(4)當地交通資訊
五、發表論文全文或摘要
本實驗室在此次國際研討會議發表的論文共兩篇，謹附於後。
Development of Microbead-based Affinity Biosensor by Insulator-Based Dielectrophoresis
Tsung-Min Chuo
1, Wensyang Hsu
1, Shih-Kang Fan
1,21Department of Mechanical Engineering, National Chiao Tung University, TAIWAN
2Department of Materials Science and Engineering, National Chiao Tung University, TAIWAN [email protected]
Abstract—This research describes a high sensitivity microfluidic bead-based immunosensor based on the principle of insulator-based dielectrophoresis (iDEP). An insulator film with small holes between two electrodes creates a nonuniform electric field. By applying appropriate voltage and frequency, the fluorescent beads are concentrated to lower electric field regions due to the difference of dielectric properties. This concentrating step enhances the fluorescence intensity of analytes and decreases the detection limit of immunosenser. In this research, the fluorescence dye is conjugated with streptavidin which has high affinity to biotin. We use biotin-labeled polystyrene beads to bind with streptavidin, therefore, we can further detect fluorescent streptavidin conjugates by a fluorescence microscope. The biotin-labeled polystyrene beads perform not only various chemical characteristics by labeling different functional groups but also offer an increased surface area for antibodies or antigens to immobilize on. Finally, we fabricate a microfluidic bead-based immunosensor with high sensitivity (1 pg/ml), short analysis time (~10 minutes), few sample consumption (~0.5 μl) and without physical microchannel.
Keywords-component; immunosensor; iDEP; fluorescence
1. Introduction
Immunoassays are biochemical analysis methods based on the high selectivity between antibody and antigen; they normally measure the presence or concentration of a specific substance in solutions or mediums that frequently contain a complex mixture of substances. Moreover, immunoassays are among the most sensitive and specific analytical methods that are routinely used in a clinical laboratory and other biological research applications.
In recent years, a new technique that uses microbeads as a solid support in immunoassays has become usual. There are several advantages in the use of microbeads. First, the microbeads’ surface to volume ratio is greater than that of a microtiter plate commonly used in conventional immunoassays.
For example, 1 g of microbeads with a diameter of 0.1μm has a total surface area of about 60 m2 [1]. The large surface area provides a large interface and the reaction field between samples and reagents. The sensitivity of immunoassays would be increased as a result of the higher efficiency of the immunoreactions between the immobilized antibody and the antigen present in a continuous flow. In addition, the reaction
Second, the immunoassays which use microbeads as a solid support can be easily integrated into a microfluidic chip. The samples and reagents that used in immunoassays can be easily transported in a fluidic system by a syringe pump or another way. Third, there are various available surface modifications for microbeads. DNA, RNA, antibodies, antigens and a vast number of other biological molecules can be easily fixed on the surface of microbeads. Moreover, transportation and analysis in a fluidic system is easy [2].
Furthermore, the dynamic condition that utilizing both diffusion and convectional forces to deliver or mix samples with reagents in microfluidic system. In contrast, conventional immunoassay on a microtiter plate, likes enzyme-linked immunosorbent assay (ELISA), is a static condition that merely depends on diffusion of the molecules for interaction and binding.
Microfluidic technology is widely used in immunoassays available to improve the analytical characteristic performances, such as short analysis time, high reliability and high detecting sensitivity, easy handling and low consumption of reagents [3].
However, a retention method is necessary for trapping or fixing microbeads in microfluidic system in order to avoid the microbeads washing away in the microfluidic system. For example, microbeads can be trapped by arrayed microstructures [4]-[6], Kitamori and coworkers fabricated a dam structure for retaining polystyrene microparticles in a glass-based microchannel [6]. Magnetic beads are also used for immunoprotein support and separation, since these beads can be easily manipulated in the channel by applying a magnetic field [7]-[9]. Dielectrophoresis [10][11], and electrostatic forces [12][13] are another way to be a retention method.
Sensitivity means the lowest concentration or the smallest amount of analytes that can be detected above the baseline, which is perhaps the most widely touted measures of an assay since it is easy quantified. Compared to conventional immunoassays, those relying on fluorescence detection, are known to be highly sensitive [14]. They have the potential that can replace the traditional ELISA technique if the fluorescence signal arising from fluorophores bound with analytes can be effectively reinforced.
In this research, we can enhance the fluorescence intensity in a simple way instead of complex chemical operations. The fluorescence intensity can be increased by concentrating beads.
We have developed a fluorescent bead-based immunoassay
bead-based immunosensor has high sensitivity, short analysis time, few sample consumption and without any microchannel.
2. Theory 2.1.
DielectrophoresisDielectrophoresis (DEP) is an electrokinetic phenomenon which can drive particles by using electrodes instead of moving actuators. A dielectric particle suspending in a solution would be affected by a force caused by the interaction between the spatially inhomogeneous electrical fields causing polarization.
The DEP force has been widely used to manipulate, transport, separate and sort different types of particles.
DEP can be classified into two types: positive DEP (p-DEP) and negative DEP (n-DEP). Particles are attracted to the region of a stronger electric field with the p-DEP force because their permittivity is greater than that of the solution. In contrast, particles are attracted to the region of a weaker electric field with the n-DEP force because their permittivity is smaller than that of the solution. In addition, p-DEP [15][16] and n-DEP [17][18] have been used to manipulate particles and biological cells with microelectrode systems.
The DEP force, FDEP, on a suspended spherical particle in a solution is given by
,
is the real part of the Clausius-Mossotti factor, given by~ ,
2.2.
Insulator-based dielectrophoresisInsulator-based (electrodeless) dielectrophoresis (iDEP) is a technology to produce the nonuniform electrical field by insulators for driving DEP. Hence iDEP would avoid the problems caused by electrodes.
In this research, iDEP is used to collect fluorescent beads on a specific device which includes two electrodes with a patterned insulator film in between as shown in Fig. 1. When voltage is applied on the device, the charged fluorescent beads are gathered at the region of weaker electric field on the insulator film, so that the beads collection can be completed by the n-DEP force. The fluorescence signal will be enhanced by
concentrating fluorescent beads. The method can increase fluorescence intensity and sensitivity. Our device is based on the iDEP technique to detect a limited amount of streptavidin.
3. Experiment 3.1.
FabricationIndium tin oxide (ITO) is one of the most widely used transparent conducting oxides because of its electrical
Indium tin oxide (ITO) is one of the most widely used transparent conducting oxides because of its electrical