microfluidics - download.e-bookshelf.de€¦ · microfluidics but also how to link these...

Microfluidics for Biological Applications

Wei-Cheng Tian Erin Finehout Editors


1 3

Editors Wei-Cheng Tian Erin Finehout General Electric General Electric Global Research Center Global Research 1 Research Circle 1 Research CircleNiskayuna NY 12309 Niskayuna NY 12309

ISBN 978-0-387-09479-3 e-ISBN 978-0-387-09480-9

Library of Congress Control Number 2008930844

copy 2008 Springer Science+Business Media LLC All rights reserved This work may not be translated or copied in whole or in part without the writtenpermission of the publisher (Springer Science+Business Media LLC 233 Spring Street New York NY 10013 USA) except for brief excerpts in connection with reviews or scholarly analysis Use in connection with any form of information storage and retrieval electronic adaptation computer software or by similar or dissimilar methodology now known or hereafter developed is forbidden The use in this publication of trade names trademarks service marks and similar terms even if they are not identified as such is not to be taken as an expression of opinion as to whether or not they are subject to proprietary rights

Printed on acid-free paper

springercom

To my family

Erin Finehout

To Gan-Wu Yu-Hsien Wei-Hua Kaitlyn Darren and Jennifer

Wei-Cheng Tian

Preface

In Nobel Prize winner Richard Feynmanrsquos well-known 1959 speech ldquoTherersquos Plenty of Room at the Bottomrdquo [1] he marvels that although many biological systems such as cells are very small they are active and perform a number of functions He then poses the challenge ldquoConsider the possibility that we too can make a thing very small which does what we want ndash that we can manufacture an object that maneuvers at that levelrdquo [1] In this book we hope to show readers that we are getting closer to meeting this challenge We have tools to manipulate and analyze small volumes of biomolecules (such as DNA and protein) we can manipulate and analyze individual cells and we can create nanodrops the size of a cell to perform specific chemical reactions All of these have been made possible by the application of microfluidics

This book consists of a selection of review articles that are intended to show how microfluidics is applied to solve biological problems why mi-crofluidics continues to play an important role in this field and what needs to be done next We will introduce not only the various technologies of microfluidics but also how to link these technologies to different biological

give perspective on the history and development of microfluidic technolo-gies They also serve to give a physical understanding of microfluidic de-vices Chapter 1 covers the physics and fluid dynamics of microscale flows Chapter 2 summarizes the materials and methods used to fabricate microfluidic devices in biological applications Chapter 3 gives solutions to how these microscale devices can be interfaced with the macro scale

have been used to study and manipulate specific classes of components Microfluidic devicessamples (Chapter 4) separate and analyze protein mixtures (Chapter 5)

specific biological applications of microfluidics tissue engineering (Chap-ter 7) high throughput screening (Chapter 8) diagnostics (Chapter 9) and

applications at the industrial and academic level Chapters 1-3

6world Chapters 4-66 give overviews on how microfluidic systems

have been used to prepare amplify and analyze DNA

and culture separate and analyze cells (Chapter 6) Chapters 7-10 focus on

biodefense (Chapter 10) And finally Chapter 11 discusses

viii

emerging trends in the microfluidics field and the current challenges to the growth and continuing success of the field

In all the chapters the authors give information on the biological problems that need to be solved the current research that is being done to address them and the obstacles that still remain In addition there are summaries of the types of products that have been commercialized in each area This book is intended to be used at the senior undergraduate or graduate level for students It will also be a great resource for researchers and scientists in the biotechnology pharmaceutical and life science industries We hope to provide the readers with an overview of microfluidics and its current ap-plications to encourage readers to think about how these technologies could help them in their own fields

Reading through the chapters there are a few recurring themes that merit being mentioned here The first is that the application of microfluidics isnrsquot just about saving time cutting costs and needing less reagents Working in the microfluidic regime enables scientists to perform experiments and use techniques that simply arenrsquot possible at a larger scale The second theme is that for the microfluidics field as a whole to continue to move forward in the biological area it is vital that scientists from different fields (engineers chemists material scientists biologist etc) work together Only with such collaborations can one be sure that the right questions are being addressed the right methods are being applied and the optimal tools are being used

The editors would like to thank Steven Elliot and Angela DePina at Springer for their help in pulling this book together We would also like to show our appreciation to the authors for all of the time and effort they put towards writing their chapters Lastly wersquod like to thank our friends and family for their support and patience during this project

References

1 Feynman RP (1960) Therersquos plenty of room at the bottom An invitation to en-ter a new field of physics Engineering and Science 2322-36

Contents

Chapter 1 Introduction to Microfluidics 1Abstract11 Introduction to Microfluidics212 History of Microfluidics 3

121 The beginning Gas chromatography and capillary electrophoresis3122 The microfluidic advantage 5123 Modular separation reaction and hybridization systems 7124 Integrated systems 8

13 Fluidics and Transport Fundamentals10131 The continuum approximation10132 Laminar flow 10133 Diffusion in microfluidic systems 12134 Surface forces and droplets14135 Pumps and valves 16136 Electrokinetics 16137 Thermal management 18

14 Device Fabrication18

x

141 Materials 19142 Fabrication and assembly 20

15 Biological Applications 21151 Genetic analysis (DNARNA) 22152 Proteomics 22153 Cellular assays 23154 Drug delivery and compatibility24

16 The Future26161 Potential demandmarket for microfluidic devices26162 Current products 27163 Challenges and the future 28

References 29

Chapter 2 Materials and Microfabrication Processes for Microfluidic Devices 35

Abstract3521 Introduction 3622 Silicon Based Materials 37

221 Micromachining of silicon39222 Bulk micromachining 39223 Surface micromachining46

23 Glass Based Materials49231 Microfabrication in glass 51

24 Wafer Bonding 56241 Fusion bonding 57242 Anodic bonding 57243 Adhesive bonding58

25 Polymers 59251 Microfabrication 59252 Polymer materials64

26 Conclusion82References 82

Chapter 3 Interfacing Microfluidic Devices with the Macro World93Abstract9331 Introduction 9432 Typical Requirements for Microfluidic Interfaces 9433 Review of Microfluidic Interfaces95

331 World-to-chip interfaces95332 Chip-to-world interfaces103

34 Future Perspectives112References 113

xi

Abstract11741 Introduction 118

411 Status of genetic analyses 118412 Genetic analysis by miniaturized electrophoresis system 119

42 Microchip Electrophoresis for Genomic Analysis122421 Material and fabrication of electrophoresis microchips 123422 Theory of gel electrophoresis of DNA 125423 Gel matrices126424 Novel DNA separation strategies on microchips130425 Surface coating methods for microchannel walls 134

43 Parallelization in Microchip Electrophoresis13744 Integration in Microchip Electrophoresis for Genetic Analysis 139

441 Sample preparation on microchip139442 System integration 141

45 Commercial Microfluidic Instruments for Genetic Analyses144451 Commercial microchip electrophoresis instruments for genetic analysis 145452 Integrated microfluidic instruments for genetic analyses 147

46 Microfluidic Markets and Future Perspectives150References 151

Chapter 5 Microfluidic Systems for Protein Separations 165Abstract16551 Introduction 166

511 Advantages of microfluidic chips for protein separations166512 Limitations of microfluidic chips in proteomics applications167513 Substrates used for proteomic analysis167

52 Microfluidic Chips for Protein Separation168521 Microchip-based electrophoretic techniques 169522 Microchip chromatography 172

53 Integrated Analysis in Microchips175531 Integration of sample preparation with analysis175532 Multi-dimensional separation in microchips 177533 Chips integrated with mass spectrometry180

54 Future Directions 180References 181

Chapter 6 Microfluidic Systems for Cellular Applications185Abstract18561 Introduction 186

611 Physiological advantages188

Chapter 4 Genetic Analysis in Miniaturized Electrophoresis Systems117

xii

612 Biological advantages189613 Economical advantages 191

62 Microfluidic Technology for Cellular Applications 191621 Microfluidic cell isolationseparation191622 Microfluidic cell culture 200623 Microfluidic cell analysis 208

63 Commercialization of Microfluidic Technology 21164 Concluding Remarks 214References 215

Chapter 7 Microfluidic Systems for Engineering Vascularized Tissue Constructs223

Abstract22471 Introduction 22472 Generating 2D Vascularized Tissue Constructs Using Microfluidic Systems22673 Generating 3D Vascularized Tissue Constructs Using Microfluidic Systems23074 Hydrogel-based Microfluidic Systems for Generating Vascularized Tissue Constructs23275 Mathematical Modeling to Optimize the Microfluidic Systems for Generating Vascularized Tissue Constructs 23576 Future Challenges 23777 Conclusions 237References 237

Chapter 8 High Throughput Screening Using Microfluidics241Abstract24181 Introduction 24282 Cell-Based Assays 244

821 High throughput cell culture245822 Cell sorting for high throughput applications252

83 Biochemical Assays254831 PCR 254832 Electrophoresis 255833 Others 255

84 Drug Screening Applications25885 Users and Developers of μF HTS Platforms 259

851 Users Research labs academic screening facilities and pharmaceutical260852 Commercialized products in HTS 261

86 Conclusion262

xiii

87 Acknowledgements 263References 263

Chapter 9 Microfluidic Diagnostic Systems for the Rapid Detection and Quantification of Pathogens 271


911 Infectious pathogens and their prevalence272912 Traditional pathogen detection methods274913 Microfluidic techniques276

92 Review of Research 277921 Pathogen detectionquantification techniques based on detecting whole cells 277922 Pathogen detectionquantification techniques based on detecting metabolites released or consumed294923 Pathogen detectionquantification through microfluidic immunoassays and nucleic acid based detection platforms297

93 Future Research Directions305References 307

Chapter 10 Microfluidic Applications in Biodefense323Abstract323101 Introduction 324102 Biodefense Monitoring 326

1021 Civilian biodefense 3261022 Military biodefense328

103 Current Biodefense Detection and Identification Methods 3301031 Laboratory detection3311032 Field detection 332

104 Microfluidic Challenges for Advanced Biodefense Detection and Identification Methods333105 Microscale Sample Preparation Methods 335

1051 Spore disruption3361052 Pre-separations 3361053 Nucleic acid purifications337

106 Immunomagnetic Separations and Immunoassays 3391061 Immunomagnetic separations 3401062 Immunoassays 341

107 Proteomic Approaches345108 Nucleic Acid Amplification and Detection Methods 346

1081 PCR and qPCR detection of pathogens for biodefense 3471082 Miniaturized and Microfluidic PCR348

xiv

1083 Heating and cooling approaches3491084 Miniaturized PCR and qPCR for biodefense3501085 Other Nucleic acid amplification methods 351

109 Microarrays3521091 Microarrays and microfluidics353

1010 Microelectrophoresis and Biodefense35410101 Microelectrophoresis technologies 356

1011 Integrated lab-on-a-chip systems and biodefense35810111 Full microfluidic integration for biodefense363

1012 Summary and Perspectives 363References 365

Chapter 11 Current and Future Trends in Microfluidics within Biotechnology Research 385

Abstract385111 The Past ndash Exciting Prospects386112 The Present ndash Kaleidoscope-like Trends 388

1121 Droplet microfluidics3891122 Integrating Active Components in Microfluidics 3911123 Third world - paper microfluidics ndash George Whitesides 3941124 Microfluidic solutions for enhancing existing biotechnology platforms3951125 Microfluidics for cell biology ndash seeing inside the cell with molecular probes 4001126 Microfluidics for cell biology ndash high throughput platforms 401

113 The Future ndash Seamless and Ubiquitous MicroTAS403References 405

Index413

List of Contributors

Beebe David J PhD University of WisconsinndashMadison Madison WI 53706 USA

Borenstein Jeffrey PhDDraper Laboratory Cambridge MA 02139 USA

Burns Mark A PhD University of Michigan Ann Arbor MI 48109 USA

Chang Dustin S University of Michigan Ann Arbor MI 48109 USA

Chang Hsueh-Chia PhDCenter for Microfluidics amp Medical Diagnostics Notre Dame IN 46556 USA University of Notre Dame Notre Dame IN 46556 USA

Chung Yao-Kuang University of Michigan Ann Arbor MI 48109 USA

xvi

Cropek Donald PhD US Army Corps of Engineers Champaign IL 61822 USA

Du Yanan PhD Massachusetts Institute of Technology Cambridge MA 02139 USA Harvard Medical School Cambridge MA 02139 USA

Gordon Jason E PhD Midwest Research Institute Kansas City MO 64110 USA

Horn Joanne PhD Microchip Biotechnologies Inc 6693 Sierra Lane Suite F Dublin CA 94568 USA

Jain Akshat University of Michigan Ann Arbor MI 48109 USA

Jovanovich Stevan PhD Microchip Biotechnologies Inc 6693 Sierra Lane Suite F Dublin CA 94568 USA

Khademhosseini Ali PhD Massachusetts Institute of Technology Cambridge MA 02139 USA Harvard Medical School Cambridge MA 02139 USA

xvii

Kuo Chuan-Hsien University of Michigan Ann Arbor MI 48109 USA

Langelier Sean M University of Michigan Ann Arbor MI 48109 USA

Lee Abraham P PhD University of California at Irvine Irvine CA 92697 USA Micronano Fluidics Fundamental Focus (MF3) Center Irvine CA 92697 USA

Lin Gisela PhD Micronano Fluidics Fundamental Focus (MF3) Center Irvine CA 92697 USA

Mofrad Mohammad R Kaazempur PhD University of California at Berkeley Berkeley CA 94720 USA

Noori Arash McMaster University Hamilton ON L8S 4L7 CANADA

Park Jihyang University of Michigan Ann Arbor MI 48109 USA

Puccinelli John P University of WisconsinndashMadison Madison WI 53706 USA

xviii

Rhee Minsoung University of Michigan Ann Arbor MI 48109 USA

Selvanganapathy P Ravi PhD McMaster University Hamilton ON L8S 4L7 CANADA

Sengupta Shramik PhD University of Missouri Columbia MO 65211 USA

Shaikh Kashan A PhD GE Global Research 1 Research Circle Niskayuna NY 12309 USA

Sommer Greg J PhD University of Michigan Ann Arbor MI 48109 USA

Takayama Shuichi PhD University of Michigan Ann Arbor MI 48109 USA

Tavana Hossein PhD University of Michigan Ann Arbor MI 48109 USA

Upadhyaya Sarvesh McMaster University Hamilton ON L8S 4L7 CANADA

xix

Wang Fang University of Michigan Ann Arbor MI 48109 USA

Wang Hong PhD Louisiana State University Baton Rouge LA 70803 USA

Wang Xuefeng PhD GE Global Research 1 Research Circle Niskayuna NY 12309 USA

Weinberg Eli J PhD Draper Laboratory Cambridge MA 02139 USA

Zeitoun Ramsey I University of Michigan Ann Arbor MI 48109 USA

Zhu Li PhD GE Global Research 1 Research Circle Niskayuna NY 12309 USA

Chapter 1 Introduction to Microfluidics

Greg J Sommer Dustin S Chang Akshat Jain Sean M Langelier Jihyang Park Minsoung Rhee Fang Wang Ramsey I Zeitoun and Mark A Burns

University of Michigan Ann Arbor MI 48109

Correspondence should be addressed to

Mark Burns (maburnsumichedu)

Keywords microfluidics history fundamentals applications commer-cialization

Abstract

Microfluidics ndash the manipulation and analysis of minute volumes of fluid - has emerged as a powerful technology with many established and relevant applications within the biological sciences Over three decades of research has yielded a wealth of techniques for improving biological assays through both the miniaturization of existing methods as well as the development of novel analytical approaches In this introductory chapter we provide an overview of microfluidic technology beginning with a historical look at the fieldrsquos origins We also present brief synopses of the fundamental physical phenomena driving microfluidics the techniques employed in de-vice fabrication and biological applications that have benefited from mi-crofluidic implementation Finally we conclude with an outlook to the fu-

2 Sommer et al

ture of the field as microfluidic technology shifts from research laborato-ries into commercial ventures This chapter is meant to familiarize readers who may be new to the field of microfluidics while highlighting areas that will be explored in more detail throughout the text

1 Introduction to Microfluidics

Microfluidic technology has evolved over the past few decades from a mo-lecular analysis endeavor aimed at enhancing separation performance through reduced dimensions into a diverse field influencing an ever-expanding range of disciplines Microfluidic techniques are being em-ployed in chemistry biology genomics proteomics pharmaceuticals bio-defense and other areas where its inherent advantages trump standard methodologies

From a biological standpoint microfluidics seems especially relevant con-sidering that most biological processes involve small-scale fluidic trans-port at some point Examples stem from molecular transfer across cellular membranes to oxygen diffusivity through the lungs to blood flow through microscale arterial networks Microfluidics can also provide more realistic in vitro environments for small-scale biological species of interest Figure 11 provides comparative length scales for several biological structures as well as common micro-fabrication structures used in microfluidic and MEMS technology

Fig 11 Approximate length scales for several biological and micro-fabrication structures

Introduction to Microfluidics 3

In this chapter we will provide a brief introduction to microfluidics begin-ning with a historical perspective of the fieldrsquos origins and concluding with an outlook on the future We also present some fundamental transport principles materials and fabrication basics and specific biological applica-tions with the aim of familiarizing the reader with important microfluidic-related concepts and highlighting areas that will be explored further throughout the text

12 History of Microfluidics

121 The beginning Gas chromatography and capillary electrophoresis

The mid-20th century saw explosive growth in the applicability of chroma-tography a technique that revolutionized the field of separation sciences by exploiting molecular distributions between mobile and stationary phases within a column Theoretical work by Golay [1] on gas chromatog-raphy (GC) and van Deemter [2] on liquid chromatography established scaling arguments showing that improved performance could be achieved by reducing open column diameters and packed column particle sizes Thus columns began being fabricated from fused silica capillaries with di-ameters on the order of micrometers Around the same time capillary elec-trophoresis (CE) was gaining popularity as a method to separate charged biomolecules Here too small bore capillaries proved advantageous as the larger surface area-to-volume ratio allowed for higher applied electric fields and therefore improved separation performance

But the roots of microfluidics truly lie in the microelectronics industry As chemists and biologists were searching for means to further miniaturize their analytical methods the microelectronics industry was improving its silicon-based micromachining processes using photolithography etching and bonding techniques [3] The merging of the bioanalytical and microe-lectronics disciplines can be considered the birth of microfluidics The first silicon-based analysis system was published in 1979 in which Terry et alfrom Stanford University fabricated a miniature GC air analyzer on a sili-con wafer [4] (Fig 12) However it was the seminal works by Manz and others in the early 1990rsquos that demonstrated the microfluidic potential for addressing issues facing analytical methods and spawned the term micro Total Analysis Systems (μTAS) [5-7]

4 Sommer et al

Fig 12 Miniature gas chromatograph developed by Terry et al on a silicon wafer in 1979 [8] - Reproduced by permission of The Royal Society of Chemistry

While researchers continued miniaturizing gas and liquid chromatography columns [6 9] many of the first successful microfluidic separation devices employed electrophoretic techniques due to the relative simplicity of ap-plying an electric potential to a microchannel versus a high-pressure source such as those required for high pressure liquid chromatography (HPLC) In 1992 Manz et al demonstrated the first on-chip CE system and initiated the new microfluidic era in separation sciences That same year Mathies et alproposed high-throughput electrophoretic sequencing on arrays of microfluidic devices [10] In 1993 Harrison et al demon-strated a micro-CE system in glass which could separate amino acids with about 75000 theoretical plates in 15 seconds [11] The next year Woolley and Mathies successfully miniaturized a microfluidic capillary gel electro-phoresis system for DNA analysis which boasted separation times in as little as 120 seconds [12]

The microfluidics boom had begun The mid 1990s brought many new concepts and devices as researchers began to investigate microfluidic uses for not only separations but other applications as well So why all the ruckus In the next section we will highlight some advantages that micro-fluidics can provide over conventional macroscale methodologies


122 The microfluidic advantage

Silicon micromachining enabled fabrication of channels and features with precision on the order of 1μm This technological feat enabled the manipu-lation of micro- (10-6) to atto- (10-18) liter volumes of fluid Such control brings several advantages from both analytical and economic viewpoints Here we briefly outline those advantages while noting that many of these concepts will be further explored throughout this chapter and text A sum-mary of the advantages are listed in Table 11

Table 11 Summary of advantages attained with microfluidic systemsMicrofluidic Advantage Description Less sample and reagent consumption

Microfluidic devices typically require 102 ndash 103

less sample volume than conventional assays

Enhanced heat transfer Higher surface area-to-volume ratio of microflu-idic channels increases effective thermal dissipa-tion

Faster separations Higher E-fields results in faster sample migra-tion

Laminar flow Low Reynolds number flows reduce sample dis-persion

Electrokinetic manipulation

Electroosmotic flow enables fluid pumping with flat plug-like velocity profiles solely via ap-plied E-fields

Lower power consumption

Fewer components and enhanced thermal dissi-pation require less power input

Parallelization Several assays can be ldquomultiplexedrdquo or run in parallel on a single chip

Portability System integration and reduced power allows for assays to be conducted using portable hand-held device

Improved separation efficiency

Efficiency in electrophoretic and chroma-tographic separations (ie number of theoretical plates) proportional to Ld

6 Sommer et al

Several measures of analytical performance can be improved through miniaturization Perhaps one of the most obvious advantages of smaller channel sizes is reduced reagent consumption leading to less waste and more efficient assays Reduced reagent consumption becomes especially advantageous for many biological applications where reagents can be very expensive (eg antibodies) and sample volumes are often limited Addi-tionally the separation efficiency (ie number of theoretical plates) of chromatographic and electrophoretic systems is proportional to Ld the length of the separation channel over its diameter Therefore long and nar-row channels enable improved peak-peak resolution Because they are so narrow microfluidic channels also boast flows with very low Reynolds numbers often Re lt 1 meaning the flow is laminar Such laminar flows inhibit additional dispersion from affecting the band width of a separated plug Diffusion however is more prominent at smaller scales and can be advantageous for mixing applications where despite very laminar flow mixing can occur solely via diffusion Narrow channels also dissipate heat more efficiently allowing for higher electric fields in electrophoretic sys-tems without adverse Joule heating effects on separation efficiency As a result the assays will require less time as higher electric fields lead to faster separations

Microfluidic devices often achieve fluid transport with few mechanical components which can significantly reduce an assayrsquos complexity and power consumption over its macroscale counterpart For example elec-troosmosis is a process in which bulk electrolytic fluid in a channel is dragged via viscosity by migrating ions near an inherently charged channel wall under the application of an electric field Electroosmosis allows for a) bulk fluid ldquopumpingrdquo using only electric fields thereby eliminating any moving parts and b) a plug-like non-parabolic fluid velocity profile that eliminates dispersion caused by parabolic pressure-driven flow Bulk fluid transport has also been demonstrated on microfluidic devices using other pumping techniques such as capillary wicking evaporation thermal gradi-ents and chemically-induced flow

With many conventional assays it is possible to integrate all analytical steps (sample loading rinsing reactions separation detection etc) into a single fully-automated platform Such integration reduces necessary hu-man involvement potential environmental contamination and analysis time With μTAS or lab-on-a-chip (LOC) devices such integration can greatly reduce the cost per analysis while providing high throughput through parallelized or multiplexed devices They can also be potentially integrated into a portable hand-held format for a variety of point-of-care


(POC) applications where proper laboratory access is not available or rapid analysis time is required including bedside patient care military and bor-der patrol and global healthcare scenarios As fabrication procedures be-come more standardized the cost per chip will decrease enabling the pro-duction of inexpensive single-use disposable chips

123 Modular separation reaction and hybridization systems

Along with the initial demonstrations of microfluidic separation systems in the early 1990rsquos researchers began exploring other methods with which to fill the ldquoanalytical toolboxrdquo necessary to build the envisioned integrated systems Further motivation for the microfluidics community arose from the explosion of genomics in the 1990rsquos Biologists were increasingly in-terested in exploring DNA and decoding genes and chromosomes as evi-denced by the appeal and success of the Human Genome Project There-fore much of the early research was directed towards DNA amplification hybridization and sequencing

One technology that naturally found its way into microfluidic devices was that of microarrays Microarrays in which minute biological samples are immobilized as individual spots that may hybridize with an introduced sample allow for extremely large numbers of parameters to be screened at any one time In 1991 the use of standard lithography to pattern an array was first introduced by Fodor et al [13] (Fig 13) Since then microfabri-cated arrays have found a home using microfluidic technologies Microar-rays have been developed to pack a maximum amount of DNA strands into a minimal amount of space As early as 1995 a 96 microwell array was used to detect an organismrsquos gene expressions [14] A year later a DNA-chip which had 1046 different DNA strands was developed demonstrating the potential power of microfabricating DNA analysis devices [15]

8 Sommer et al

Fig 13 DNA microarray fabricated lithographically by Fodor et al in 1991 Each square represents one gene sequence and is 50μm wide (Reprinted from [13] with permission from AAAS)

plications Aside from relatively simple reactions within microarrays an important biological reaction that benefited from miniaturized formats is the polymerase chain reaction (PCR) PCR is a technique that uses enzy-matic transcription to systematically amplify parts or all of a DNA strand triggered via thermal fluctuations On-chip PCR incorporation was made possible due to rapid and efficient heating and cooling of extremely small sample volumes allowing for quick and proficient thermocycling In 1995 Northrup et al developed the first microfabricated device capable of ther-mocycling and PCR reactions [16] On-chip PCR would prove to be in-strumental in future DNA sequencing and genotyping devices

124 Integrated systems

Modular system integration is particularly advantageous in μTAS devices for several reasons First the sample can be isolated from the outside envi-ronment reducing error caused from human contact and sample contami-nation In addition having all processes located on a single chip can in theory reduce sample-processing time and allow for a fully-automated analysis This potential has prompted the idea of integrated devices for point-of-care diagnostics and clinical analysis However full integration of numerous modular components is not a trivial task and requires the resolu-tion of numerous problems including sample injection pumping data dis-semination and product retrieval

Early microfluidic reactors were also primarily focused on biological ap-


At the onset of modular device development integrated systems were gen-erally simple and relatively crude Technological efforts were focused on designing simplistic systems that performed one or two actions instead of many One of the first integrated DNA analysis systems was presented by Burns et al in 1998 (Fig 14) This device was engineered to meter a pre-cise DNA sample volume mix it with reagents amplify it separate it us-ing gel electrophoresis and detect the fluorescence signal on-chip [17] Since that time DNA separation stages have also been equipped with both on-chip fluorescent and conductivity detectors for on-chip analysis How-ever a problem plaguing many of these systems from a diagnostic stand-point is that they lack an easy and reliable way to process raw sample such as blood or saliva

Fig 14 Integrated DNA analysis device developed by Burns et al in 1998 (Re-printed from [17] with permission from AAAS)

During its infancy microfluidics exhibited the potential to explode into a field rich with powerful applications and commercial successes Some lik-ened its promising impact to that of the integrated circuit (IC) industry Today however the microfluidics field is arguably not where it was envi-sioned ten years ago So what factors have impeded to the fieldrsquos emer-gence as a prevailing technology We will further explore that question later on in the chapter but for now we remind the reader that microfluidics continues to be a vibrant and active research area with researchers tackling the challenges hindering its widespread acceptance

10 Sommer et al

13 Fluidics and Transport Fundamentals

Researchers employ many fundamental transport and scaling principles for manipulating and analyzing microfluidic flows In this section we intro-duce some of the important concepts governing microscale transport with the hope of elucidating several advantages of microfluidics over macro-scale techniques We also introduce several dimensionless groups (Table 12) that can be used to evaluate the relative importance of different phe-nomena

131 The continuum approximation

Unlike solids fluids (especially gases) consist of molecules that are rea-sonably widely separated However in fluid mechanics despite the fact that properties like velocity and density vary wildly at the molecular scale we usually view fluids as ldquocontinuousrdquo and discuss ldquoaveragerdquo fluid proper-ties rather than considering the properties of each molecule Does this con-tinuum approximation still hold in microfluidics where fluid volumes are very small

Indeed for the volumes typically encountered in microfluidics the contin-uum approximation remains sufficiently valid A 1 picoliter [(10 μm)3] volume of fluid still contains 3 x 1013 water molecules large enough for us to consider their average rather than individual behavior Typically for most properties the continuum approximation does not break down until we approach length scales on the order of several molecular diameters [18]

The continuum approximation is important because it allows us to analyze microfluidic flows with the same governing principles developed for mac-roscale fluid mechanics However as researchers continue to drive fluids into smaller length scales (ie nanofluidics and beyond) the continuum approximation starts to break down and we must develop new approaches for analyzing these small regimes

132 Laminar flow

Microfluidic devices almost always boast smooth laminar flow as opposed to turbulent flow which is of a stochastic nature and marked by the pres-ence of ldquoeddiesrdquo that disrupt parallel streamlines The Reynolds number


(Re) is a dimensionless parameter used to determine the transition from laminar to turbulent regimes with Re lt 2100 considered laminar for flow in cylindrical channels The Reynolds number for this flow is defined as

ν

ud=Re

(11)

where u is the flow velocity ν is the kinematic viscosity and d is the channel diameter In fluid mechanics terms the Reynolds number com-pares the magnitudes of inertial force to viscous forces in a flow Because

dpropRe the small dimensions of microfluidic channels are responsible for very low Reynolds numbers resulting in laminar flows In fact for most microfluidic applications Re lt 1

Since microfluidic flows are usually laminar simple flows like Poiseuille flow are commonly encountered Poiseuille flow occurs when we have steady fully-developed pressure driven flow of a Newtonian fluid in a channel The velocity profile for Poiseuille flow is parabolic with the maximum velocity being in the center of the channel The equations for Poiseuille flow in a cylindrical channel are as follows [19]

])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)

where vz is the velocity of the fluid along the axis U is the mean velocity (maximum velocity at centerline of channel = 2U) R is the radius of the channel r is the radial distance from the axis z is the distance along the axis P is the pressure along the channel μ is the fluid viscosity and Q is the volumetric flow rate

While laminar flow is advantageous in many microfluidic applications (ie in electrophoretic separations for reducing band broadening due to disper-sion or in diffusion-based separation devices such as the H-filter) it can also be a nuisance such as in processes where mixing is necessary Most chemical and biological assays require mixing or dilution at some point

12 Sommer et al

Several clever techniques have been developed to achieve mixing in lami-nar regimes mostly through geometric design or by taking advantage of enhanced diffusivity across small dimensions

133 Diffusion in microfluidic systems

Diffusion ndash the stochastic process by which molecules drift from one re-gion to another ndash is another property that takes on increasing importance in microfluidic systems as channel dimensions are reduced Diffusive trans-port is driven by random thermal motion of particles such that given enough time and the absence of external influences a species will be ho-mogeneously distributed throughout a finite volume

Most microfluidic systems combine diffusive transport with convective flow of the bulk fluid therefore it is helpful to compare the relative impor-tance of each effect The Sherwood number a dimensionless number rep-resenting the ratio of convective mass transfer to diffusive mass transfer in a system is defined as

Sh =kd

D(15)

where k is the mass transfer coefficient d is the characteristic length of the system (eg channel diameter) and D is the diffusion coefficient For most macroscale systems Sh is large resulting in convective transport being dominant over diffusive transport However for microfluidic systems the Sherwood number is much lower due to the presence of the characteristic length scale in the numerator Therefore diffusion assumes much more importance in microfluidic systems

Diffusive transport without flow (or perpendicular to streamlines with flow) can be estimated from the simplified mass transfer equation

2

2

x

CD

t

C

part

part=

part

part (16)

Consider a stationary liquid system with a step change in species concen-tration such that the concentration is held at C for all time t at a certain location Let there be no other source for the substance in the liquid and no other mode of transport besides diffusion At time t the concentration will

be C2 at a distance of about Dt from the source and will be 1 of C


at a distance of about 4 Dt (Fig 15) Therefore diffusion has a signifi-

cant effect up to a distance of about 4 Dt For polymers or proteins in a fluid D is on the order of ~10-7 cm2s So for a time period of about 10 seconds the distance at which diffusion is significant will be about 40μm a length that is usually negligible for macroscale purposes but can be a significant distance in microfluidic systems Note that at twice that dis-

tance (ie 8 Dt ) the concentration is only 10-8C

Fig 15 Diffusion-dependent concentration profile at time t for a solute with diffu-sion coefficient D

Several mixing and separation techniques have been developed exploiting this diffusive effect in microfluidic systems Readers are especially re-ferred to the H-filter [20] developed by Paul Yagerrsquos lab at the University of Washington for an example of an efficient and highly-effective device that uses this very simple concept (Fig 16) Here species in a stream are separated into two diverging flows based on their differing diffusivities across a microchannel

Wei-Cheng Tian Erin Finehout Editors


1 3


ISBN 978-0-387-09479-3 e-ISBN 978-0-387-09480-9




springercom

To my family

Erin Finehout


Wei-Cheng Tian

Preface











viii





References


Contents





x




References 29











xi
















xii











86 Conclusion262

xiii















xiv










Index413








xvi








xvii









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)










ISBN 978-0-387-09479-3 e-ISBN 978-0-387-09480-9




springercom

To my family

Erin Finehout


Wei-Cheng Tian

Preface











viii





References


Contents





x




References 29











xi
















xii











86 Conclusion262

xiii















xiv










Index413








xvi








xvii









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









To my family

Erin Finehout


Wei-Cheng Tian

Preface











viii





References


Contents





x




References 29











xi
















xii











86 Conclusion262

xiii















xiv










Index413








xvi








xvii









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









Preface











viii





References


Contents





x




References 29











xi
















xii











86 Conclusion262

xiii















xiv










Index413








xvi








xvii









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









viii





References


Contents





x




References 29











xi
















xii











86 Conclusion262

xiii















xiv










Index413








xvi








xvii









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









Contents





x




References 29











xi
















xii











86 Conclusion262

xiii















xiv










Index413








xvi








xvii









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









x




References 29











xi
















xii











86 Conclusion262

xiii















xiv










Index413








xvi








xvii









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









xi
















xii











86 Conclusion262

xiii















xiv










Index413








xvi








xvii









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









xii











86 Conclusion262

xiii















xiv










Index413








xvi








xvii









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









xiii















xiv










Index413








xvi








xvii









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









xiv










Index413








xvi








xvii









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)
















xvi








xvii









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









xvii









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









xviii









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









xix













Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)















Abstract


2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









2 Sommer et al












4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)















4 Sommer et al





















6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)


























6 Sommer et al









8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)














8 Sommer et al










10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)













10 Sommer et al







132 Laminar flow




ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)











ν

ud=Re

(11)




])(1[2)( 2

R

rUrvz minus=

(12)

U = minusR2

8μ

dP

dz

(13)

dz

dPRQ

μ

π

8

4

minus=(14)



12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









12 Sommer et al





Sh =kd

D(15)



2

2

x

CD

t

C

part

part=

part

part (16)









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