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1、第1-4章 微流控芯片,目 录,发展背景制备技术流体控制典型实例商品简介,目 录,发展背景制备技术流体控制典型实例商品简介,Do you know?,Biochip生物芯片Lab-on-a-Chip 芯片实验室LabchipMicrofluidic Chip 微流控芯片Micro Total Analysis System(MicroTAS,TAS)微全分析系统,Research into miniaturization is primarily driven by the need to reduce costs by reducing the consumption of expensiv
2、e reagents and by increasing throughput and automation.For example,most are aware of the increasing cost of health care,driven in part by the cost of implementing the latest diagnostic assays.These assays,which are usually performed in microtiter plates that consume hundreds of microliters of reagen
3、ts,would benefit from the use of microfabricated arrays of nanoliter volume vials.By reducing reagent consumption by a factor of 103104,these devices could provide dramatic savings for the repetitive assays often performed in diagnostic laboratories.,Why miniaturization？,Anal.Chem.2000,72,330A-335A,
4、In the same way that integrated circuits allowed for the miniaturization of computers from the size of a room to the size of a notebook,miniaturization has the potential to shrink a room full of instruments into a compact lab-on-a-chip.,Anal.Chem.2000,72,330A-335A,尺寸效应（cm100m）尺寸1/100分子扩散时间1/10,000(1
5、h0.36s)体积1/1,000,000试剂用量1t1g,mlnl传热速度1500oC/s由此将带来：方法上的变革理论上的突破还有巨大的经济和社会效益,微流控分析芯片,微流控分析芯片目的是通过化学分析设备的微型化与集成化，最大限度地把分析实验室的功能转移到便携的芯片中。微流控分析芯片通过微机电加工技术把整个实验室的功能，包括采样、稀释、加试剂、反应、分离、检测等集成在几平方厘米的微流控芯片上，且可多次使用，因而极大地减少了样品和分析试剂的用量，降低了分析的成本，加快了分析的速度，具有广泛的适用性。,The Early Days:1975-1989,The first analytical mi
6、niaturized deviceA gas chromatographic air analyzer fabricated on a silicon wafer Terry,S.C.Ph.D.Thesis,Stanford,Stanford,CA,1975Terry,Stephen C.;et al.IEEE Transactions on Electron Devices,1979,ED-26(12),1880A miniature gas anal.system based on the principles of gas chromatog.The major components a
7、re fabricated in Si using photolithog.and chem.etching techniques,which allows size redns.of nearly 3 orders of magnitude compared to conventional laboratory instruments.consists of a sample injection valve,a 1.5-m-long capillary column.A thermal conductivity detector fabricated on a separate silico
8、n wafer.sepns.of gaseous hydrocarbon mixts.are performed in 10 s.,发 展 历 史,Photograph of a gas chromatograph integrated on a planar silicon wafer fabricated by Terry and co-workers at Stanford University.,However,the response of the scientific community to this first silicon chip device was virtually
9、 none,presumably because of the lack of technological experience(of the separation scientists)to deal with this kind of device.the research work related to miniaturization on silicon focused on the fabrication of components such as micropumps,microvalves,and chemical sensors.,The Renaissance:1990-19
10、93,the reemergence of silicon-based analyzersDesign of an open-tubular column liquid chromatograph using silicon chip technologyManz,A.;et al.Sensors and Actuators,B:Chemical(1990),B1(1-6),249 A novel concept of high pressure liquid chromatog.a silicon chip with an open-tubular column and a conducto
11、metric detector.A 55mm chip containing an open-tubular column of 6 m2m15cm was fabricated,which has theor.separation efficiencies of 8000 and 25,000 plates in 1 and 5 min,resp.The total column volume is 1.5 nL and the detection cell volume 1.2 pL.,Micrograph of Liquid Chromatograph chip manufactured
12、 by Manz and co-workers at Hitachi Ltd.,The concept of miniaturized total chemical analysis system or TAS was proposed by Manz et al.the main reason for miniaturization was therefore to enhance the analytical performance of the device rather than to reduce its size.it was also recognized that a smal
13、l size presented the advantage of a smaller consumption of carrier,reagent,and mobile phase.,Growing to Critical Mass:1994-1997,In 1994,the number of published papers related to TAS increased abruptly since more research groups joined the efforts to develop the area.Microfabrication Design Separatio
14、ns Biochemical Reactors Detection,分类与特点,分类：材料：硅、玻璃、石英、聚合物、复合材料功能：分离、采样与前处理、检测、化学合成等特点：高效、低耗、集成、一致性好、昂贵,目 录,发展背景制备技术流体控制典型实例商品简介,微结构的形成1经典的光刻技术,Photolithograph procedures for making glass template.(a)Spine coating of photoresist,(b)covered with photo mask,(c)exposure,(d)developing,(e)etching,and(f)re
15、moval of photoresist.,适合硅、玻璃、石英等材料，与传统的半导体工业的方法一致。分为湿法和干法两种，干法的分辨率较湿法高，相应的制造成本也高。,Analyst,2004,129,305308,制备技术 之,微结构的形成2模版浇注法（模塑法）,Process overview for mass manufacturing of plastic microfluidic systems,适合聚合物材料。大批量生产时成本低。,Anal.Chem.,2002,74,78A-86A,微结构的形成3模版热压法,Schematic representation of the fabric
16、ation method involving hotembossing of thermoplastic polymer pellets and thermal bonding.,适合热塑性聚合物。,Applied Physics Letters,2002,80,3614-3616,微结构的形成4激光刻蚀法,用激光直接在聚合物或玻璃上加热形成微结构.,Anal.Chem.,1997,69,2035-2042,Microfilter,Sensors and Actuators B 67 2000 203208,芯片的封装1热键合,对玻璃和石英材质刻蚀的微结构一般使用热键合方法，将加工好的基片和相
17、同材质的盖片洗净烘干对齐紧贴后平放在高温炉中，在基片和盖片上下方各放一块抛光过的石墨板，在上面的石墨板上再压一块重0.5kg的不锈钢块，在高温炉中加热键合。玻璃芯片键合时，高温炉升温速度为10oC/分，在620oC时保温3.5小时，再以10oC/分的速率降温。石英芯片键合温度高达1000oC以上。此方法对操作技术要求较高。,现代科学仪器，2001，4，8-12,制备技术 之,芯片的封装2阳极键合,在玻璃、石英与硅片的封接中已广泛采用阳极键合的方法。即在键合过程中，施加电场，使键合温度低于软化点温度。在500-760伏电场下，升温到500oC时，可使两块玻璃片键合。在两块玻璃板尚未键合时，板间空
18、气间隙承担了大部分电压降，玻璃板可视为平行板电容器，板间吸引力与电场强度的平方成正比，因此，键合从两块玻璃中那些最接近的点开始，下板中可移动的正电荷（主要是Na+）与上板中的负电荷中和，生成一层氧化物（正是这层过渡层，使两块玻璃板封接），该点完成键合后，周围的空气间隙相应变薄，电场力增大，从而键合扩散开来，直至整块密合。,现代科学仪器，2001，4，8-12,芯片的封装3室温键合,Anal.Chem.2004,76,5597-5602,芯片的封装4贴合 将聚合物薄片直接覆盖在玻璃或石英板上。5压合,Schematic illustration of sealing and connection
19、 method.The top and bottom plates are pressed by a screw and holders.,Anal.Chem.2002,74,1724-1728,目 录,发展背景制备技术流体控制典型实例商品简介,遵循低雷诺数流动的规律。除了组分间的扩散，两层或者多层流体可以相邻流动而不互相混合，使得样品的混合变得困难。,Anal.Chem.2002,74,45-51,液体流动的特点,液体流动的特点,由于比表面积增大，表面张力、摩擦力的影响非常显著。,微通道中的液液界面与通道壁平行，因为表面张力和摩擦力大于重力。,Anal.Sci.2001,17,89-93,液
20、体流动的特点,液体的物理性质发生变化，如表观粘度变大,纯水通过微通道的时间：理论值2.3ms实验值10ms,Anal.Chem.2002,74,6170-6176,Microfabrication InsideCapillaries Using MultiphaseLaminar Flow Patterning,Science 1999,285,83,液体流动的特点,Microchip Flow Cytometry Using Electrokinetic Focusing,Anal.Chem.1999,71,4173,液体流动的特点,A Picoliter-Volume Mixer for
21、Microfluidic Analytical Systems,Anal.Chem.2001,73,1942-1947,液体的混合,Ultrasonic Mixing in Microfluidic Channels Using Integrated Transducers,Anal.Chem.,2004;76;3694-3698,液体的混合,外加压力驱动,Anal.Chem.2000,72,1711-1714,重力驱动,Anal.Chem.2005,77,1330-1337,离心力驱动,Centrifugal Microfluidics Platform,Anal.Chem.2002,74,
22、5569-5575,毛细作用驱动,Microfilter device design and detail:(a)top view of generic device design with narrow and expanded channels,(b)filter detail area showing filter pores and expanded channel layout,(c)microfilter cross section.,LabChip,2005,5,922-929,单向阀,Cylinder diameters are approximately 100 and 25
23、m.,Anal.Chem.2002,74,4913-4918,目 录,发展背景制备技术流体控制典型实例商品简介,Schematic illustration of experimental setup and ion pair extraction model,Anal.Chem.2001,73,1382-1386,离子检测,determination of Co(II)as 2-nitroso-1-naphthol chelates by solvent extraction and thermal lens microscopy,Lab Chip 2001,1,72-75,Schemati
24、c diagrams of CE procedures with pinched injection(top panels),floating injection(middle),and(C)simplest injection mode(bottom).,Anal.Chem.,2001.73,2656-2662,电泳分离,General idea of polymer membrane formation under organic/aqueous two-phase flow in an X-shaped microchannel,Anal.Chem.2003,75,350-354,固定化
25、酶反应,Substrate permeation and subsequent enzyme reaction experiment,Anal.Chem.2003,75,350-354,Schematic illustrations of microchip-based immunosorbent assay.,Anal.Chem.2001,73,1213-1218,免疫检测,Glass microchip for immunosorbent assay:,Design of a Compact Disk-like Microfluidic Platform for Enzyme-Linked
26、 Immunosorbent Assay,Schematics of(a)a CD-ELISA design with 24 sets of assays,(b)a single assay,(1,waste;2,detection;3,first antibody;4,6,8,10,washing;5,blocking protein;7,antigen/sample;9,second antibody;and 11,substrate),and(c)photo of a single assay.,Anal.Chem.,2004,76(7),1832-1837,免疫检测,(a)Schema
27、tic of five-step flow sequencing CD(1,waste;2,detection;7,antigen/sample;8,9,washing;10,second antibody;and 11,substrate)and(b)a CNC-machined CD.,Nature Biotechnology，2008，26，1373-1378,免疫检测,Anal.Chem.2004,76,1824-1831,核酸分析,Self-Contained,Fully Integrated Biochip for Sample Preparation,Polymerase Cha
28、in Reaction Amplification,and DNA Microarray Detection,Microfabricated Device for DNA and RNA Amplification by Continuous-Flow Polymerase Chain Reaction and Reverse Transcription-Polymerase Chain Reaction with Cycle Number Selection,Anal.Chem.2003,75,288-295,PCR反应,Development of a Microchip-Based Bi
29、oassay System Using Cultured Cells,Concept of the microchip-based bioassay system,Anal.Chem.,2005.77(7),2125-2131,细胞培养和操作,(A)Illustration and(B)photo showing the arrangement of Peltier elements.(C)Photo of the temperature control device,(A)Illustration and(B)photograph of the bioassay microchip.(C)A
30、n enlarged illustration of the microchamber for cell culture,Complete system for the microchip-based bioassay,(A)Photo of the temperature control device with the microchip.(B)Thermograph of the microchip surface,Micrographs of(A)mouse peritoneal macrophages and(B,C)J774.1 cells in the microchip.The
31、dam successfully kept cells in the microchamber.,Micrographs of J774.1 cells cultured in the microchip for 2 days(A)under continuous medium flow conditions and(B)static conditions.The cells stained in blue were dead.,Chemical processes carried out in the microchip for bioassay of macrophage-stimulat
32、ing agent,Calibration curves of NO dissolved in the medium(A)determined on the bulk scale and(B)in a microchip.,Typical results of NO released from J774.1 cells.,目 录,发展背景制备技术流体控制典型实例商品简介,微流控芯片主要应用相关商业化产品,微流控芯片供应链情况,Caliper-LabChip Systems,LabChip GX,LabChip XT,LabChip EZ Reader,LabChip XT分馏系统进行快速，自动
33、化分离核酸,LabChip GXII是一个蛋白质样品的精确的分析完整的解决方案，是一种蛋白质研究用户的理想工具。利用微流体技术，无需处理SDS-PAGE凝胶。,LabChip EZ Reader是一个突破性的台式仪器，有利于激酶检测和其他酶的筛选和分析。以微流控分离为基础。,GE Healthcare-BiacoreSystems,Serial flow cell systems,Independentflow cell systems,Agilent-1200 HPLC Chip,Agilent-2100 Bioanalyzer,安捷伦2100芯片生物分析仪是目前最为成功的基于微流控的商业化
34、平台，提供用于DNA、RNA、蛋白质和细胞分析的解决方案，30 分钟内给出结果，提供自动化的、高品质的数字化数据。安捷伦 2100 生物分析仪平台产品系列包括：标准安捷伦 2100 生物分析仪除了电泳分析外，还支持芯片上的流式细胞术，并符合21CFR第11部分的要求。,世界上第一个整合全自动样品制备和检测程序的实时定量PCR仪。直接加入采样样品,GeneXpert自动完成样品裂解、核酸纯化浓缩、定量PCR扩增检测，并输出分析结果，整个过程只需要30分钟！,microfluidic cartridges for sample preparation,Cepheid-Genexpert,Raind
35、ance-RDT 100,Droplets are processed on a disposable chip without moving parts or valves for robust operation,RDT 1000采用Rainstorm微滴为基础的技术扩增数百成千上万的基因位点具有较高的特异性和一致性。RDT 1000最大限度地减少错误，同时避免多重PCR技术的复杂性，利用这一行之有效的方法的灵敏度和特异性PCR的效率最大化。,Fluidigm-BioMark System,BioMark高通量基因分析系统是集成微流控芯片技术、实时定量PCR技术及强大的基因分析软件相结合的
36、技术平台。其创新技术就在于集成液体通路技术：利用集成电路制作工艺（光刻）在硅片或石英玻璃上刻上许多微管和微腔体，通过不同的控制阀门控制溶液在其中的流动来实现生物样品的分液、混合、PCR扩增。集成流体通路技术极大地简化了生物样品和试剂的分液操作，提高分析通量和灵敏度，其纳升级的反应体系为高通量的基因分析应用节省大量成本,Fluidigm-EP1 System,EP1系统提供了最有效的高通量SNP基因分型。它允许极低的运行成本，并为低到中多重SNP基因分型提供最简单的工作流程。EP1系统也是世界上第一个可重复使用的SNP基因分型芯片。,Cellectricon-Dynaflow HT,优越的离子通道筛选系统 允许任何离子通道的目标的 高效筛选 全自动-8小时无人操作 降低运行成本的10倍,
