第八屆全國微全分析系統學術會議微全分析專場

　　2013年5月16日-19日，由中國化學會主辦、廈門大學承辦、復旦大學、浙江大學協辦的為期四天的第八屆全國微全分析系統學術會議、第三屆全國微納尺度生物分離分析學術會議暨第五屆國際微化學與微系統學術會議在美麗的海濱城市廈門隆重召開。以下是微全分析專場報告。

上海交通大學 任吉存教授

　　來自上海交通大學的任吉存教授為我們帶來了題為《Microfluidic Chip with Fluorescence Correlation Spectroscopy》的精彩報告，以下是內容摘要：

　　In this work, we presented an analytical system for coupling fluorescence correlation spectroscopy (FCS) with microfluidic chip (as shown in Figure 1). FCS is a sensitive single molecule technique based on measuring the fluorescence fluctuations in a small volume. When a molecule diffuses into and out of the tiny illuminated volume, or when a chemical reaction occurs in this volume, a fluorescence fluctuation can be tracked. The highly-focused detection volume was obtained with a high numeric aperture (NA) water immersion objective. The photon signals were collected by a single photon counting module. Utilizing the water immersion objective with 60×1.2 (NA) and the 35 μm pinhole, the highly-focused Gaussian volume was obtained of about 0.3 fL. Meanwhile, we developed a simple method for fabrication of poly(dimethylsiloxane) (PDMS)/glass microfluidic chip[2]. An irreversible sealing was formed between the PDMS replica and glass by UV irradiation. This microfluidic chip with FCS was used to capillary zone electrophoresis and isotachophoresis. This method was used to measure the surface charge of quantum dots and determine the circulating DNA in sera samples (as shown in Figure 2).

浙江大學 牟穎教授

　　來自浙江大學的牟穎教授為我們帶來了題為《自吸離散化數字PCR 芯片的研究》的精彩報告，以下是內容摘要：

　　數字PCR 技術能進行單分子擴增，實現核酸初始拷貝數絕對定量，因此被認為是極為準確靈敏的檢測技術[1]，在生命科學領域意義重大。目前報道的主要數字PCR 裝置有：離心轉盤芯片[2]、滑動芯片[3]、液滴芯片[4，5]以及集成流路(IFC)芯片[6]。然而上述裝置需要繁雜的外部設備，如注射泵、氣壓源等控制系統來實現樣品的注入以及分配操作，進樣耗時長，操作繁瑣，易造成樣品的丟失，降低檢測的靈敏性及準確性。本文研制了一種樣品自吸式無泵無閥數字PCR 芯片，該裝置應用PDMS 本身所具有的容氣性質，靠儲存的負壓引入樣品，通過油相封隔的方法自動實現樣品在各個小室的分配，實現了數字PCR 進樣和隔離的自動化，通過PCR 方法實現核酸的單分子擴增檢測，如圖所示。基于自吸式離散化芯片的數字PCR 不僅具有和目前幾類商品化的數字PCR 平臺同樣的準確性(絕對定量)和敏感度(單分子)，而且更為簡便、省時，容易推廣應用。 

中山大學 吳天準教授

　　來自中山大學的吳天準教授為我們帶來了題為《基于潤濕特性仿生的PDMS 功能涂層自圖形化》的報告，以下是內容摘要：

　　PDMS 微流控芯片常需要各種表面改性，而已有的各種氣相、液相涂層方法都難以保證涂層覆蓋準確性和均勻性。受非洲沙漠甲蟲及超親水葉子的潤濕特性啟發，我們提出一種極為高效、便捷的PDMS 表面涂層自圖形化 (self-patterning) 方法。動植物的親水、疏水潤濕特性可高效地操控液體的黏、滑行為(圖a)，因此，我們通過粗糙度設計潤濕梯度，以蒸發為驅動力便可精確地將溶質沉積、固化在粗糙的3D 微通道上，而不會留在光滑頂面(圖b)。利用氧氣等離子體選擇性攻擊SU-8 而非Si 的特點，可便捷地把SU-8 與Si 的粗糙度復制到PDMS 上，從而使涂層蒸發后只分布在通道內，兼容PDMS-玻璃邦定(圖c)。利用此方法，我們示范了疏水CYTOP 涂層在PDMS 復雜流路中的100%覆蓋準確性和良好的均勻性(圖d-e)。與無涂層PDMS 相比，溶脹被顯著抑制，非特異性熒光染料吸附被有效消除(圖f-g)。這種自圖形化方法廉價、高效、快速，可廣泛用于PDMS 微流控芯片使用各種功能涂層進行表面改性。

大連化物所 劉顯明教授

　　來自大連化物所的劉顯明教授為我們帶來了題為《Measurement and Intelligent Control of Electrowetting-actuated Droplets by Means of Electronic Approaches》的精彩報告，以下是內容摘要：

　　In digital microfluidics (DMF), transportation of droplets was determined by the on-off output (0 or 1) of its control system, which applies a voltage on the actuation electrodes underneath and thereby generates unidirectional electrowetting forces on droplets. Such an actuation mechanism provides a technical shortcut to the programmability of automation functions and even more advanced intelligent control, the latter of which autonomously coordinates objects in accordance to human intelligence and experiences.

　　Here we present a way of establishing an intelligent control-engaged actuation and routing system for droplets on DMF chips. First, we looked into the intricacy of a digital droplet in their hydrodynamics by real-time impedance measurement [1] and image capture; second, based on the data obtained from hydrodynamic studies, we used a fuzzy control module to coordinate the charging timing of working electrodes and an overall 20% shorter actuating time was obtained, compared with that engaged in a proportional-integral-derivative controller (PID controller) [2]; third, we established a routing coordinator for multiple digital droplets while providing a possibility for an autonomous and preliminary “fire-and-forget” management function.

南京大學 吳增強教授

　　來自南京大學的吳增強教授為我們帶來了題為《β-Galactosidase / Glucose Oxidase Cascade Reaction in Microfluidic system: Convection and Diffusion Interaction》的報告，以下是內容摘要：

　　In the processes of metabolism, the high energy conversion and great turnover number are exemplified by sequential enzymes catalytic reactions. These phenomena indicate multi-enzymes cascade reactions have more advantages than single enzyme in the bio-fuel cell, detections fields and modeling reaction pathways within cells1-3, However, the challenge is critical to reveal how biology integrates temporal and spatial information to determine the biological function. What is more, understanding the effect of spatial organization on enzymatic activity in multienzyme systems is not only fundamentally interesting, but also important for translating biochemical pathways to noncellular environments. Here, based on the our pervious research by using Scanning Electrochemical Microscopy (SECM)4, a study of distance-dependence for the activity of β-Galactosidase (β-Gal)/ Glucose Oxidase (GOD) cascade by immobilizing β-Gal/GOD on Polycarbonate (PC)/PDMS hybrid microchip is reported in this paper. A schematic representation of enzymes cascade reaction in microchannel is outlined in Figure 1. The influence of spatial distance and velocity on the kinetic of enzymes cascade reaction was investigated by using electrochemical detection at the end of microchip. A 2D finite element model was applied to understand the effect of mass transport on the spacing-dependent enzymes cascade reaction kinetics.

中國科學院化學研究所 陳義教授

　　來自中國科學院化學研究所的陳義為我們帶來了題為《光子晶體微流控芯片的制備與應用》的報告，以下是內容摘要：

　　微納流控方法有顯著的理論優勢，其基礎研究也因此迅速發展直至今日，唯實用化研究頗有難度。原因甚多，其中流體調控困難和通道表面非特異吸附放大效應，十分關鍵。現有芯片分離速度雖快，但峰容量過小，因而無力分辨復雜樣品中的諸多成分，這是一個更現實的問題，至今難解。我們對此進行了一些探索，考慮了引入穩流和峰擴容機制，提出了構造規則或周期性的微納復合通道結構的策略，嘗試了將光子晶體[1]Photonic crystals，記為PCs)融入微流控概念的做法，設計制備出了PC 型微納流控芯片，研究了其高速分離問題并取得了進展。PCs 是一類周期性電介質物質，既有天然產物，亦可人工合成，在光學通訊、光子線路設計、光學器件研制等領域中的發展非常迅速，但在分離分析領域中的研究進展緩慢。我們認為，利用PCs 中巨大的比表面積和3D 納米孔網結構，可以誘導高穩流動，獲得高速和高效分離，因為納米通孔不僅具有尺寸篩分作用，還能縮短傳質距離，強化物質的分配作用。實測表明，PC 型微納流控芯片的分離塔板高度可小于300nm，優于普通毛細管電泳;此類芯片不僅可以實現高效、高速、高重現性的分離[2]，而且可以有效提高峰容量，能用于混合氨基酸(圖1)、肽或蛋白水解產物等復雜樣品的分離分析;結合使用手性選擇劑，還可實現手性物質的高速拆分。種種跡象顯示，PC 型微納流控芯片深具發展潛力，值得大力研究。不過，目前還沒有商品可用，須自己制備。我們已為此建立了快速制備PCs 的方法[2]，可用于不同形狀和結構芯片中高穩定性PCs 的構建，歡迎查考引用。

南京大學 徐靜娟教授

　　來自南京大學徐靜娟教授為我們帶來了題為《微流控芯片上的電致化學發光分析》的報告，以下是內容摘要：

　　微流控芯片的信號檢測裝置是微流控分析檢測系統的重要組成部分。檢測器的性能對整個分析系統的靈敏度、線性范圍、檢測速度有很大的影響。電致化學發光(ECL)法結合了電化學與化學發光的優勢，因此，它有著很高的靈敏度與選擇性。隨著雙極電極的出現，微流控芯片在ECL 方面的研究也越來越多。它與傳統的三電極系統有著明顯的不同：雙極電極上的電位由溶液中的電場決定的，而且不需要用引線將電極引出，這大大的簡化了芯片制作的復雜程度。到目前為止這種方法還很少被應用于分析檢測領域。我們實驗室用光刻法和化學刻蝕法制備了光透的ITO 雙極電極，由于ITO 電極的透光性使得它非常適合于高靈敏的ECL 檢測。利用葉酸與腫瘤細胞表面高表達的葉酸受體之間的高親和能力以及葉酸對釕聯吡啶的高效猝滅效果實現對腫瘤細胞的檢測，并觀察到了葉酸在細胞內的胞吞胞吐現象。除了檢測細胞表面物質，這種無線電致化學發光法還首次用于細胞內的c-Myc mRNA 的檢測。由于mRNA 容易水解，因此，我們用耐水解的DNA 取代mRNA。首先將雙鏈DNA 用量子點標記，再轉移到細胞內。通過DNA 與mRNA 的雜交，使得蛋白質的翻譯受阻，引起腫瘤細胞的凋亡。同時使得單鏈DNA 從量子點上釋放，并被電極表面的釕聯吡啶納米粒子標記的DNA 所捕獲。無線電致化學發光技術除了用于高靈敏的化學傳感還可以用于生物成像分析。通常生物成像技術受背景信號的干擾較大，我們設計了一種雙管道的微芯片-ECL 傳感器，利用電子開關來降低背景信號，提高成像的靈敏度，成功檢測了前列腺抗原(PSA)。

廈門大學 周勇亮教授

　　來自廈門大學的周勇亮教授為我們帶來了題為《Low-cost fabrication of glass based microfluidic chips by PAG electrochemical soft stamping》的精彩報告。以下是內容摘要：

　　Glass chip is an important and commonly used materials for microchips due to its excellent properties. Conventional method for the galss chips is based on UV lithography, which requires specialliazed equipments and represnents a barrier to the mass production of glass microchips[1]. Campbell et al. proposed the direct printing microstructures onto glass by agarose stamps soaked in HF etching soluton[2]. Hanners developed a method that combines PDMS microcontact printing and wet etching[3]. However, compared with chemical etching, electrochemical etching has higher accuracy and controllability. Our group has developed electrochemical wet stamping technique, to electrochemical etch silicon or metal surface using agarose gel soft stamps[4]. Here, we reported that by using polycarylamide hydrogel (PAG) as electrochemical soft stamps, galss chips could be fabricated in mass with cost as almost one fifth of that of UV lithography.

　　The apparatus and process of electrochemical soft stamping was shown in figure 1. Electric field was applied between PAG soft stamp and the metal film waiting for fabricated. PAG soft stamp was cast on silicon/photoresist mold which was fabricated by nanoimprint, the dimension of smallest structure is around 300 nm.

　　As shown in the figure 2, the Young’s modulus of PAG stamps were obtained by fitting the approaching curve with Hertz model based on the AFM force-distance curves of the PAG stamps. The effect of the Acr and bis concentration to stamp modulus were discussed as a basis to explore the stamp stress and strain under pressure by using ANSYS simulation, the results show that the Young’s modulus of PAG gel is 4.46 MPa, which is around 2 orders of magnitude higher than that of agarose gel.

　　After optimized the processes and parameters, such as etching potential, pressure, time, pattern shape and the wetability of the surface of the fabricating metal, microgroove structures with high quality have been fabricated (Fig 3). And the fabbrication cost of glass chips is about 33.4 RMB per chip ( yield > 50), while that of UV lithography is 148RMB (Fig 4).

湖南農業大學 周鐵安教授

　　來自湖南農業大學的周鐵安教授為我們帶來了題為《Novel Voltage Controlled Solid State Pumps Using Microchannel Plates》的報告。以下是內容摘要：

　　Microchannel plates (MCP), 2.1 cm in diameter, 0.40 mm thick, and with 3 million uniform holes 7 microns in diameter, have been used for pumping ethanol, 2-propanol, methanol, acetone and water. Pumping started at dc voltages as low as 6 V, and a pumping rate of 15-18 ml/min was achieved at 200-250 V for ethanol. The pumping rate was found to be directly proportional to the voltage drop, DV, applied across the channel plate at voltages ≥ 100 V, in agreement with the conventional electro-osmotic equation. Highly reproducible results were given by several MCPs continuously working in ethanol for over one week. Enhancing pumping was realized by stacking of several MCPs in various configurations. In addition to the surface charge induced by surface dissociation and specific adsorption, an additional space charge governed by the field effect in the n-type MCP semiconductor device was found to play an important role in the electro-osmotic pumping. This new mechanism also explains well for the intriguing phenomena described in a recent publication[1] that MCP shows a unique characteristic of directional preference in pumping with net flow consistently induced toward the nearest electrode regardless of polarity of the voltage potential applied. Other electrohydrodynamic (EHD) mechanisms involved in preconditioning of the plates are also discussed. The devices have potential application in voltage controlled micropumps.

中科院上海微系統與信息技術研究所 李剛教授

　　來自中科院上海微系統與信息技術研究所的李剛教授為我們帶來了題為《A self-contained metering and mixing microfluidic device for lab on a chip》的精彩報告，以下是報告摘要：

　　Over the past decade, there has been an increased interest in the development of miniaturized analysis devices. One of the most expected features in these miniaturized devices is the integration and automation of liquid operations for biochemical analysis applications [1]. In spite of intensive effort on microfluidics, there has been no practical solution for the integrated and easy-to-use liquid handling technique. Thus, a power-free device is proposed in the present study for nanoliter fluidic handling on a chip.

　　Fig. 1 shows the schematic of the microfluidic device based on the pump/microchip assembly. It is assembled by reversibly bonding a PDMS slab containing a group of microchambers on a PDMS chip containing a microchannel network. The PDMS chip contains 24 parallel microstructure units. All units are connected by a common feeding channel and each unit comprises a centre metering structure, a peripheral metering structure, a reaction chamber, a peripheral inlet port, and an air venting port. In the combination, the PDMS slab acts as a pump to provide pumping power, which is pre-degassed and then attached to the outlet ports of the microchip to create a negative pressure for driving fluid by absorbing the air in the channels [2]. In addition, a sudden contract of cross-section is designed at the end of each metering channel, which acts as a geometrical stop-valve. During operation, a pre-degassed modular PDMS pump was firstly placed onto the outlet ports of the microfluidic chip, and then aliquots of liquid samples were dispensed into the inlet reservoirs. Due to negative pressure generated inside microchannels by the air dissolution into the pre-evacuated PDMS pump, the samples were automatically sucked into the feeding channels. When the advancing liquid fronts reached the joints between the metering channel and the connection channel, they stopped owing to capillary pressure barriers. Subsequently, air was introduced into the feeding channel to remove the excess liquid, and droplets of precise volumes were left in the metering channels. With a further absorption of the air in the closed channel-reservoir system, the negative pressure increased and pushed the metered droplets into the connection channel, and finally transported the mixed droplet into the reaction chamber. Fig. 2 shows snapshots of the metering and mixing of two food dye solutions in the proposed microfluidic device.