使用數位液滴介電潤濕技術結合類比流體開發整合型篩選收集之微型裝置

I.本篇研究為開發一個整合型以電潤濕為基礎的篩選蒐集裝置，用來篩選具有螢光性質的樣品，本實驗設計為結合毛細管連續微流道與介電潤濕技術之整合型裝置。電潤濕晶片使用鍍有氧化銦錫的導電玻璃，排除以往傳統的電潤濕晶片的設計使用非共平面 (dual planar)的方式，本實驗是使用控制電極及接地電極為共平面 (co-planar) 的方式對液滴進行操作，利用標準黃光微影製程製作控制電極 (control electrode)及接地電極 (ground electrode) 於同一晶片上，並塗佈鐵氟龍同時作為介電層及疏水層；藉由驅動注射幫浦將欲分析溶液注射於毛細管中，液滴滴落在晶片上後會同時接觸控制電極與接地電極，接著使用LabVIEW軟體搭配DAQ卡控制繼電器之切換順序進而控制晶片上的電極驅動順序切換，使液滴進行移動。在晶片實驗中，施加約150 Vrms的交流電壓 (1 kHz) 後，即能驅使液滴移動。當流道中注入螢光顆粒並使用光電倍增管搭配電壓訊號擷取卡，從LabVIEW軟體的圖表中確認PMT可準確計數通過偵測視窗的每一顆含有螢光顆粒的水滴並驅動電極將之移動至蒐集區；反之，未含有螢光顆粒的水滴則被驅動至廢液區。實驗結果證明此整合型裝置可以成功的判別含有螢光顆粒水滴及無螢光顆粒水滴之差異，並成功的篩選出兩者且準確度高。 II.實驗室自開發之離子風誘導漩渦濃縮裝置將其應用於濃縮及偵測被鍵結上拉曼探針的細菌。以離子風誘導之微型渦流將液體中的懸浮微粒聚集濃縮是一項新興微流體技術。當針尖狀電極架設8 mm直徑的容槽上方並施加高頻高壓電場，電極會將其周圍的空氣粒子極化帶電，帶電粒子累積於電極尖端達飽和時會因同性電相斥使帶電粒子排出形成離子風 (Ionic wind)。離子風吹拂在下方容槽中的液體表面會產生一道剪切作用力並推動液體表面轉動，進一步帶動下方容槽中整體液體一起旋轉產生漩渦，液體中的懸浮粒子被漩渦帶往容槽中央並聚集。研究中將利用聚苯乙烯球 (Polystyrene beads) 進行離子風濃縮系統的測試。進一步的研究中，我們嘗試使用離子風濃縮器搭配金奈米聚集團拉曼標籤 (Nanoaggregate-embedded beads, NAEB) 的技術與表面有修飾抗原之 PS 微粒子與 antibody@NAEB 進行接合，最後經過離子風系統濃縮後再以拉曼光譜儀對濃縮點進行偵測，觀察離子風是否可以順利地將 NAEB@PS 進行濃縮。本研究中的最後部分是利用離子風濃縮系統對常見於引發食物中毒的沙門氏菌 (Salmonella) 及淋病常見奈瑟氏菌 (Neisseria) 進行濃縮，由結果中發現，離子風系統能夠在短時間內將修飾上拉曼探針 NAEB 的細菌濃縮，發現離子風系統能應用於不同種細菌的濃縮上。亦發現使用拉曼訊號來進行細菌濃度的定量是可行的。

關鍵字

介電潤濕；數位液滴；螢光顆粒；光電倍增管；實驗室晶片；離子風；細菌；沙門氏菌；奈瑟氏菌

並列摘要

I. This work reports the integration efforts of capillary microchannel with the device of electro-wetting on dielectrics (EWOD). Using standard lithography processes, control electrodes are deposited on indium tin oxide (ITO) glass substrates, where Teflon film is then coated to work as dielectric and hydrophobic layer to develop the EWOD chip. The liquid is driven into the capillary using a syringe pump to form droplets hanging at the capillary outlet, which is 2.5 mm above the chip. When one drop falls on the chip to trigger the relays controlled by computer interfacing cards, the electrodes are activated in sequence to move the droplet. LabVIEW software package is used to develop controlling programs. In our experiments, the 1 kHz AC voltage (150 Vrms) is applied to drive the droplets. When one droplet containing fluorescence particles falls on the EWOD chip, the fluorescence signals detected with photomultiplier tube are sent to control cards to switch the relays and divert the droplet to collection well. Otherwise, the non-fluorescent droplets are moved to waste well. The results indicate the particle sorting processes using this EWOD device are feasible with adequate accuracy. II.Using micro- vortex driven by ionic wind generated near a corona needle tip above the liquid level of a small reservoir to provide centrifugal flows has been an emerging micro-fluidic technique to trap and concentrate suspended microparticles such as bacteria to detect. The surrounding air near the corona needle is ionized when high ac voltage is applied. The accumulated gas ions are repelled from each other at the needle tip to eject and produce ionic wind, which can swipe across the liquid air interface inside a miniaturized reservoir to generate centrifugal vortices, when the neelde is off the reservoir center. Suspended particles are further dragged into the reservoir center, where the vortex is more intense to trap at the reservoir bottom. In this work, antigen-functionalized polystyrene micro-beads were conjugated with nanoaggregate-embedded beads (NAEB), made of silica-coated gold nano-particle aggregates doped with Raman reporter molecules, and immobilized with antibody probes to demonstrate particle concentrations at the stagnant point at the reservoir bottom by ionic wind-driven centrifugal flows in a reservoir containing 60 μL solution. Clear Raman scattering signals enhanced by gold nanoparticle plasmoncis were acquired from the particle concentrated spot to identify the trapped particles. The ionic wind flows were also use to trap and concentrate Neisseria and Salmonella bacteria docked with antibody-functionalized NAEB at the level of 106 colony forming units (CFU) per ml or lower, when NAEB were doped with ethyl violet and fluorescein derivative, respectively. In addition to recognize concentrated bacteria by obtaining the Raman spectrum from the reporter molecules in NAEB, the linear relation between Raman scattering signal intensity and bacteria concentration was found from the concentration spot of NAEB-docked Salmonella over the range of 104 to 106 CFU per mL. This relation indicates the feasibility to quantify bacteria sample concentrations using this micro-centrifugal method.

並列關鍵字

Electro-wetting on dielectric (EWOD) ； Digital droplet ； fluorescent particle ； Photomultiplier ； Lab on a chip ； Ionic wind ； bacteria ； Salmonella ； Neisseria

參考文獻

1. van den Berg, A. and T. Lammerink, Micro total analysis systems: microfluidic aspects, integration concept and applications, in Microsystem technology in chemistry and life science. 1998, Springer. p. 21-49.

2. Manz, A., N. Graber, and H.á. Widmer, Miniaturized total chemical analysis systems: a novel concept for chemical sensing. Sensors and actuators B: Chemical, 1990. 1(1): p. 244-248.

3. Neuzil, P., J. Pipper, and T.M. Hsieh, Disposable real-time microPCR device: lab-on-a-chip at a low cost. Molecular bioSystems, 2006. 2(6-7): p. 292-298.

4. Watts, P. and S.J. Haswell, The application of micro reactors for organic synthesis. Chemical Society Reviews, 2005. 34(3): p. 235-246.

5. Pirone, D.M. and C.S. Chen, Using lab-on-a-chip Technologies to Understand Cellular Mechanotransduction, in Lab-on-Chips for Cellomics. 2004, Springer. p. 171-196.

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使用數位液滴介電潤濕技術結合類比流體開發整合型篩選收集之微型裝置

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