DOCSLIB.ORG

Droplet Microfluidics for Synthetic Biology

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Droplet Microfluidics for Synthetic Biology Lawrence Berkeley National Laboratory Recent Work Title Droplet microfluidics for synthetic biology. Permalink https://escholarship.org/uc/item/6cr3k0v5 Journal Lab on a chip, 17(20) ISSN 1473-0197 Authors Gach, Philip C Iwai, Kosuke Kim, Peter W et al. Publication Date 2017-10-01 DOI 10.1039/c7lc00576h Peer reviewed eScholarship.org Powered by the California Digital Library University of California Lab on a Chip View Article Online CRITICAL REVIEW View Journal | View Issue Droplet microfluidics for synthetic biology Cite this: Lab Chip,2017,17,3388 Philip C. Gach,*ab Kosuke Iwai,ab Peter W. Kim,ab Nathan J. Hillsonacde and Anup K. Singh *ab Synthetic biology is an interdisciplinary field that aims to engineer biological systems for useful purposes. Organism engineering often requires the optimization of individual genes and/or entire biological pathways (consisting of multiple genes). Advances in DNA sequencing and synthesis have recently begun to enable the possibility of evaluating thousands of gene variants and hundreds of thousands of gene combinations. However, such large-scale optimization experiments remain cost-prohibitive to researchers following tradi- tional molecular biology practices, which are frequently labor-intensive and suffer from poor reproducibil- ity. Liquid handling robotics may reduce labor and improve reproducibility, but are themselves expensive Received 30th May 2017, and thus inaccessible to most researchers. Microfluidic platforms offer a lower entry price point alternative Accepted 9th August 2017 to robotics, and maintain high throughput and reproducibility while further reducing operating costs through diminished reagent volume requirements. Droplet microfluidics have shown exceptional promise DOI: 10.1039/c7lc00576h for synthetic biology experiments, including DNA assembly, transformation/transfection, culturing, cell rsc.li/loc sorting, phenotypic assays, artificial cells and genetic circuits. Introduction transfection. The resulting derivative organisms are then ana- lyzed for the detection and quantification of desired products Synthetic biology exploits engineering principles and innova- (and/or other relevant proteins/RNAs/metabolites/etc.). Fig. 1 tions in molecular biology, cell biology, and bioinformatics to shows the key steps involved in the synthetic biology process design and construct modified organisms with desired prop- for the selection of highest-performing pathway amongst erties. Synthetic biology has demonstrated significant poten- many possible configurations, the successful implementation tial, for example, in the biomanufacturing of a wide variety of of which demands significant expertise and high-throughput commercially-viable products including pharmaceuticals, bio- instrumentation for biological design, manufacture, and per- fuels, chemicals, and biomaterials.1 Primary techniques to ge- formance screening. Most experiments are performed manu- netically modify organisms can be grouped into two distinct ally and are very labor-intensive, consume large amounts of Published on 10 August 2017. Downloaded by Lawrence Berkeley National Laboratory 23/10/2017 18:29:10. categories: directed evolution and rational design. Directed expensive reagents such as enzymes and synthetic DNA, are evolution uses randomized mutagenesis to generate diverse limited in throughput, and have poor reproducibility. Robotic DNA libraries through a variety of approaches such as gene liquid-handling stations can overcome the throughput and re- shuffling, error-prone PCR, and the use of chemical mutagens producibility limitations, however they are very expensive, or irradiation. Conversely, rational design (potentially leverag- hard to maintain, and consume the same amount of reagents ing biological part characterization data and modeling/com- as manual experiments (Table 1). puter-aided design tools) implements a set of specific genetic Microfluidic systems overcome many of the drawbacks of variants through targeted DNA synthesis/assembly/editing ap- both manual and robotic systems, as they are capable of high proaches. With either technique, exogenous constructs (if throughput, low reagent consumption, and automation. any) are introduced into host organisms by transformation/ Droplet-based microfluidics, in which sub-microliters to picoliters of aqueous phase are encapsulated into monodis- perse droplets, are especially useful for applications requiring a Technology Division, DOE Joint BioEnergy Institute, Emeryville, California parallel experiments at minimal reagent costs. In addition, 94608, USA b Biological and Engineering Sciences, Sandia National Laboratories, Livermore, droplet-based microfluidic technologies enable high- California 94550, USA. E-mail: [email protected], [email protected]; throughput and controlled processes for cell-free, artificial Fax: +1 925 294 3020; Tel: +1 925 294 1260 cells, and genetic circuits applications. Previous review arti- c Fuels Synthesis Division, DOE Joint BioEnergy Institute, Emeryville, California cles have covered the fundamentals of droplet-based micro- 94608, USA d Biological Systems and Engineering Division, Lawrence Berkeley National Lab, fluidics, droplets-in-flow technologies for biological assays, Berkeley, California 94720, USA and flow-based microfluidic methodologies for synthetic 2–4 e DOE Joint Genome Institute, Walnut Creek, California 94598, USA biology. This review focuses on recent advances in droplet- 3388 | Lab Chip,2017,17, 3388–3400 This journal is © The Royal Society of Chemistry 2017 View Article Online Lab on a Chip Critical review Fig. 1 Molecular biology and analytical steps involved in synthetic biology using droplet microfluidic systems. Biological design/build/test engineering cycles include key steps such as DNA synthesis, DNA assembly, DNA transformation, cell culture, and phenotypic analysis, which often require costly and labor-intensive manual processes. Microfluidic systems have the potential to overcome such drawbacks, through enabling high- throughput automated processing with low reagent consumption requirements. Table 1 Comparison of conventional and microfluidic approaches to synthetic biology Synthetic biology steps Conventional high throughput method Droplet microfluidics approach Metabolic pathway design - Biological computer-aided design -N/A and manufacture tools (bioCAD/CAM)121 - Sequence and design repositories de novo oligonucleotide - Controlled pore glass (CPG) column - Not available Published on 10 August 2017. Downloaded by Lawrence Berkeley National Laboratory 23/10/2017 18:29:10. synthesis - Microarray with printing technology122 Gene assembly - Sequential assembly protocol in an automated - Digital microfluidics (MDF)-based multi-step droplet robotic system123 merger10,11,55 - Microarray with micro-wells22,124,125 - Droplet-in-flow merger Transformation - Microtiter plate heat shock and electroporation - DMF-based electroporation and heat shock49 - Automated transformation plating - Droplet-in-flow-based electroporation and heat shock Outgrowth/culture - Microtiter plate incubation in robotic systems - Off-device surfactant-stabilized droplet incubation - On-device incubation - Streaking droplets on an agar plate62 Colony picking -Colony picking robots - Fluorescence-activated droplet sorter (FADS)3,69 - DMF-based sorter - Passive droplet traps Phenotypic assay - Microtiter plate reader - FADS3,69 - Mass spectrometry (MALDI-MS, LC-MS, etc.) - Droplet-in-flow to mass spectrometry18,80,81 - Fluorescence-activated cell sorting (FACS) - DMF to mass spectrometry19 Cell-free and artificial - Vesicle bioreactors126 - Flow focusing and microfluidic jetting for formation cell systems and genetic - Manual formation of bilayer of vesicles16,92 circuits - Observation of genetic circuits as - Droplet interface bilayers95 a cell population in solution or on plates - Droplets on microarray surface15 - Encapsulation of genetic circuits in droplets115,119 This journal is © The Royal Society of Chemistry 2017 Lab Chip,2017,17, 3388–3400 | 3389 View Article Online Critical review Lab on a Chip based microfluidic systems directed explicitly towards syn- molecules are soluble in them, and hence, can be lost from the thetic biology applications, which may become the fastest droplet interior. Most droplet microfluidics rely on fluorocar- growing segment of the worldwide synthetic biology market.5 bon oils and fluorocarbon surfactants, as these are biocompati- Fundamental microfluidics advances, and applications to sys- ble and have low solubility for hydrophobic molecules. Fluoro- tems biology, sequencing, single-cell analysis, and drug carbon oils also have a high gas solubility, required to support screening, (that merely have the potential for future applica- cell culture. While fluorocarbon surfactants have proven effec- tion to synthetic biology) have been purposefully excluded. tive for molecular biology steps and cell culture, they are not compatible with mass spectrometry (for example, the otherwise Droplet microfluidic formats preferred surfactant PicoSurf1 is incompatible with mass spectrometry).18,19 For mass spectrometry applications, A variety of microfluidic systems, each having inherent advan- nanoparticle-based surfactants could be a potential option.20 For tages and disadvantages, support synthetic biology applica- artificial cell applications, phospholipid is typically used instead, tions spanning DNA assembly to single-cell phenotyping. in an organic phase such as chloroform, hexane, and octanol.21
Load more

Recommended publications
  • Digital Microfluidic Devices: the Role of the Dielectric Layer
    Vasco Alexandre Violante Rodrigues Licenciatura em Ciências de Engenharia em Micro e Nanotecnologias Digital Microfluidic devices: the role of the dielectric layer Dissertação para obtenção do Grau de Mestre em Engenharia de Micro e Nanotecnologias Orientador: Professor Doutor Rui Igreja, Dep. Ciência dos Materiais, FCT-UNL Co -orientador: Professor Doutor Hugo Águas, Dep. Ciência dos Materiais, FCT-UNL Júri: Presidente: Prof. Doutor Rodrigo Ferrão de Paiva Martins Arguente: Doutora Rita Maria Mourão Salazar Branquinho Vogal: Prof. Doutor Rui Alberto Garção Barreira do Nascimento Igreja Novembro, 2014 DIGITAL MICROFLUIDICS: THE ROLE OF THE DIELECTRIC LAYER © Vasco Alexandre Violante Rodrigues Faculdade de Ciências e Tecnologia Universidade Nova de Lisboa A Faculdade de Ciências e Tecnologia e a Universidade Nova de Lisboa têm o direito, perpétuo e sem limites geográficos, de arquivar e publicar esta dissertação através de exemplares impressos reproduzidos em papel ou de forma digital, ou por qualquer outro meio conhecido ou que venha a ser inventado, e de a divulgar através de repositórios científicos e de admitir a sua cópia e distribuição com objectivos educacionais ou de investigação, não comerciais, desde que seja dado crédito ao autor e editor. AGRADECIMENTOS É com enorme satisfação que vejo cumprida mais uma etapa da minha vida, tendo sido esta porventura uma das mais exigentes até ao momento. A conclusão deste trabalho encerra em si um capítulo que começou por ser caracterizado por grande incerteza, tendo eu pertencido ao reduzido grupo de pessoas que ingressou pela primeira vez no Mestrado Integrado em Engenharia de Micro e Nanotecnologias, e é com alegria que pertenço ao grupo ainda mais restrito de pessoas que o conclui agora, cinco anos volvidos.
    [Show full text]
  • Recent Advances in Electrowetting Microdroplet Technologies
    Chapter 4 Recent Advances in Electrowetting Microdroplet Technologies Robert W. Barber and David R. Emerson 4.1 Introduction The ability to handle small volumes of liquid, typically from a few picolitres (pL) to a few microlitres (mL), has the potential to improve the performance of complex chemical and biological assays [1–6]. The benefits of miniaturisation are well documented [7–9] and include smaller sample requirements, reduced reagent con- sumption, decreased analysis time, lower power consumption, lower costs per assay and higher levels of throughput and automation. In addition, miniaturisation may offer enhanced functionality that cannot be achieved in conventional macroscale devices, such as the ability to combine sample collection, analyte extraction, preconcentration, filtration and sample analysis onto a single microfluidic chip [10]. Microfluidics-based lab-on-a-chip devices have received considerable attention in recent years, and are expected to revolutionise clinical diagnostics, DNA and protein analysis and many other laboratory procedures involving molecular biology [11–13]. Currently, most lab-on-a-chip devices employ continuous fluid flow through closed microchannels. However, an alternative approach, which is rapidly gaining popularity, is to mani- pulate the liquid in the form of discrete, unit-sized microdroplets—an approach which is often referred to as digital microfluidics [14–18]. The use of droplet-based microfluidics offers many benefits over continuous flow systems; one of the main advantages is that the reaction or assay can be performed sequentially in a similar manner to traditional (macroscale) laboratory techniques. Consequently, a wide range of established chemical and biological protocols can be transferred to the micro-scale without the need to establish continuous flow protocols for the same reaction.
    [Show full text]
  • Recent Advances in Droplet-Based Microfluidic Technologies
    micromachines Review Recent Advances in Droplet-based Microfluidic Technologies for Biochemistry and Molecular Biology 1, 1, 2, Joel Sánchez Barea y , Juhwa Lee y and Dong-Ku Kang * 1 Department of Chemistry, Incheon National University, Incheon 22012, Korea; [email protected] (J.S.B.); [email protected] (J.L.) 2 Department of Chemistry, Research Institute of Basic Sciences, Incheon National University, Incheon 22012, Korea * Correspondence: [email protected]; Tel.: +82-32-835-8094 These authors contribute equally to this article. y Received: 2 May 2019; Accepted: 18 June 2019; Published: 20 June 2019 Abstract: Recently, droplet-based microfluidic systems have been widely used in various biochemical and molecular biological assays. Since this platform technique allows manipulation of large amounts of data and also provides absolute accuracy in comparison to conventional bioanalytical approaches, over the last decade a range of basic biochemical and molecular biological operations have been transferred to drop-based microfluidic formats. In this review, we introduce recent advances and examples of droplet-based microfluidic techniques that have been applied in biochemistry and molecular biology research including genomics, proteomics and cellomics. Their advantages and weaknesses in various applications are also comprehensively discussed here. The purpose of this review is to provide a new point of view and current status in droplet-based microfluidics to biochemists and molecular biologists. We hope that this review will accelerate communications between researchers who are working in droplet-based microfluidics, biochemistry and molecular biology. Keywords: droplet-based microfluidic; biochemistry; molecular biology; digital PCR; biochip; biosensor; digital quantification; microfluidic; single cell analysis 1.
    [Show full text]
  • Detection of Dna Hybridization on a Configurable Digital Microfluidic
    DETECTION OF DNA HYBRIDIZATION ON A CONFIGURABLE DIGITAL MICROFLUIDIC BIO- CHIP USING SPR IMAGING Lidija Malic1, Teodor Veres2 and Maryam Tabrizian1 1Biomedical Engineering Department, McGill University, CANADA and 2Industrial Materials Institute, National Research Council, CANADA ABSTRACT This paper presents a configurable digital microfluidic-based surface plasmon resonance (SPR) biochip platform comprising an electrowetting-on-dielectric (EWOD) microfluidic device coupled to SPR imaging (SPRi). We demonstrate its application for dynamic on-chip simultaneous immobilization of different DNA probes in combination with multichannel label-free real-time detection of subse- quent hybridization reactions. The integrated EWOD-SPRi system would enable the development of high-throughput, rapid and ultrasensitive biomolecular detection strategies beyond DNA microarray applications. KEYWORDS: Digital microfluidics, EWOD, SPR imaging, DNA hybridization INTRODUCTION EWOD microfluidics has attracted considerable attention in the past decade and the most recent efforts are directed towards its application in biomedical research [1- 3]. While these studies demonstrate the versatility of EWOD devices, the reported applications involve homogeneous phase reactions [4] and detection methods that require labeled biomolecules [2] or sample extraction from the chip [1]. This in- creases both the time and complexity of the assay. To introduce new applications relying on label-free, real-time surface sensitive detection techniques such as SPRi, it would be advantageous to use droplet-based EWOD actuation for surface specific biomolecule immobilization. However, the need of hydrophobic properties for EWOD actuation renders immobilization of biomolecules such as DNA on the sur- face of the chip impossible [3, 4]. In this paper, we demonstrate for the first time the use of an EWOD microfluidic chip to dynamically immobilize DNA probes in a two-dimensional array, followed by SPRi detection of bioaffinity interactions.
    [Show full text]
  • Interfacing to Biological Systems Using Microfluidics
    University of Tennessee, Knoxville TRACE: Tennessee Research and Creative Exchange Doctoral Dissertations Graduate School 12-2018 Interfacing to Biological Systems Using Microfluidics Peter Golden Shankles University of Tennessee, [email protected] Follow this and additional works at: https://trace.tennessee.edu/utk_graddiss Recommended Citation Shankles, Peter Golden, "Interfacing to Biological Systems Using Microfluidics. " PhD diss., University of Tennessee, 2018. https://trace.tennessee.edu/utk_graddiss/5315 This Dissertation is brought to you for free and open access by the Graduate School at TRACE: Tennessee Research and Creative Exchange. It has been accepted for inclusion in Doctoral Dissertations by an authorized administrator of TRACE: Tennessee Research and Creative Exchange. For more information, please contact [email protected]. To the Graduate Council: I am submitting herewith a dissertation written by Peter Golden Shankles entitled "Interfacing to Biological Systems Using Microfluidics." I have examined the final electronic copy of this dissertation for form and content and recommend that it be accepted in partial fulfillment of the requirements for the degree of Doctor of Philosophy, with a major in Energy Science and Engineering. Scott T. Retterer, Major Professor We have read this dissertation and recommend its acceptance: Steven M. Abel, Mitchel J. Doctycz, Jennifer L. Morrell-Falvey Accepted for the Council: Dixie L. Thompson Vice Provost and Dean of the Graduate School (Original signatures are on file with official studentecor r ds.) Interfacing to Biological Systems Using Microfluidics A Dissertation Presented for the Doctor of Philosophy Degree The University of Tennessee, Knoxville Peter Golden Shankles December 2018 Copyright © 2018 by Peter Golden Shankles All rights reserved.
    [Show full text]
  • Demonstration of Automated DNA Assembly on a Digital Microfluidic Device
    Rochester Institute of Technology RIT Scholar Works Theses 5-12-2021 Demonstration of Automated DNA Assembly on a Digital Microfluidic Device Hee Tae An [email protected] Follow this and additional works at: https://scholarworks.rit.edu/theses Recommended Citation An, Hee Tae, "Demonstration of Automated DNA Assembly on a Digital Microfluidic Device" (2021). Thesis. Rochester Institute of Technology. Accessed from This Thesis is brought to you for free and open access by RIT Scholar Works. It has been accepted for inclusion in Theses by an authorized administrator of RIT Scholar Works. For more information, please contact [email protected]. ROCHESTER INSTITUTE OF TECHNOLOGY ROCHESTER, NY Demonstration of Automated DNA Assembly on a Digital Microfluidic Device by Hee Tae An A Thesis Submitted in Partial Fulfillment of the Requirements for the Degree of Master of Science in Microelectronic Engineering Submitted May 12, 2021 DEPARTMENT OF ELECTRICAL AND MICROELECTRONIC ENGINEERING KATE GLEASON COLLEGE OF ENGINEERING Approved By: Dr. Michael Schertzer, Associate Professor Date Thesis Advisor, Department of Mechanical Engineering Dr. Karl Hirschman, Professor Date Committee Member, Department of Electrical and Microelectronic Engineering Dr. Ivan Puchades, Assistant Professor Date Committee Member, Department of Electrical and Microelectronic Engineering Dr. Michael Schrlau, Associate Professor Date Committee Member, Department of Mechanical Engineering Dr. Sean Rommel, Director of Microelectronic Engineering/Professor Date Department Representative, Department of Electrical and Microelectronic Engineering Abstract The rapid manufacturing of highly accurate synthetic DNA is crucial for its use as a molec- ular tool, the understanding and engineering of regulatory elements, protein engineering, genetic refactoring, engineered genetic networks and metabolic pathways, and whole-genome syntheses [1,2].
    [Show full text]
  • Protein Stamping for Maldi Mass Spectrometry Using an Electrowetting-Based Microfluidic Platform
    PROTEIN STAMPING FOR MALDI MASS SPECTROMETRY USING AN ELECTROWETTING-BASED MICROFLUIDIC PLATFORM Vijay Srinivasan1, Vamsee Pamula, Phil Paik, and Richard Fair Department of Electrical and Computer Engineering, Duke University, Durham, NC 27708-0291. ABSTRACT MALDI-MS (matrix-assisted laser desorption/ionization mass spectrometry) is one of the most commonly used tech- niques for protein analysis. In conventional systems sample preparation is typically done in well-plates and transferred onto a MALDI target by robotic systems, which are complex, huge, expensive and slow. In this paper, we present a droplet-based microfluidic interface to transfer protein samples from a well-plate format onto a MALDI target for MS analysis. The droplets are actuated using the electrowetting phenomenon, and are immersed in silicone oil which pre- vents non-specific adsorption and enables the manipulation of high concentrations of proteins. Droplet transport and droplet formation were evaluated as a function of protein concentration using bovine serum albumin (BSA) as a test system. Droplet transport was possible for BSA concentrations up to 10mg/mL which is three orders of magnitude higher than previously reported results on handling proteins by electrowetting. Droplet formation from on-chip reser- voirs, using only electrowetting forces and no external pressure assistance, was possible up to concentrations of 0.01mg/mL. An interface between a well-plate format and the electrowetting chip, and a scheme to passively stamp droplets onto a target substrate was then designed and tested by stamping BSA solutions. In two separate experiments 3.6fmoles and 16fmoles of BSA were stamped onto a glass slide using 0.001mg/mL and 0.01mg/mL samples respec- tively.
    [Show full text]
  • Interfacing Microfluidics to LDI-MS by Automatic Robotic Spotting
    Microfluid Nanofluid (2010) 8:777–787 DOI 10.1007/s10404-009-0510-x RESEARCH PAPER Interfacing microfluidics to LDI-MS by automatic robotic spotting Chia-Wen Tsao • Song Tao • Chien-Fu Chen • Jikun Liu • Don L. DeVoe Received: 30 July 2009 / Accepted: 23 September 2009 / Published online: 13 October 2009 Ó Springer-Verlag 2009 Abstract We developed a method of interfacing micro- Keywords Mass spectrometry interfacing Á fluidics with mass spectrometry (MS) using a robotic Microfluidics Á Robotic spotting Á LDI-MS spotting system to automate the contact spotting process. We demonstrate that direct and automated spotting of analyte from multichannel microfluidic chips to a custom 1 Introduction microstructured MALDI target plate was a simple, robust, and high-throughput method for interfacing parallel The ability to perform high-throughput bioanalysis in a microchannels using matrix-assisted laser desorption/ioni- single integrated platform is a substantial benefit offered by zation mass spectrometry (MALDI-MS). Using thermo- microfluidics technology. Arrays of multiplexed micro- plastic cyclic olefin copolymer (COC) polymer microfluidic channels allowing large numbers of simultaneous on-chip chips containing eight parallel 100 lm 9 46 lm micro- analyses have been demonstrated in numerous applications channels connected to a single input port, spotting volume including chemical reaction screening (Wang et al. 2006a), repeatability and MALDI-MS signal uniformity are evalu- immunoassays (Wang et al. 2006b), enzymatic processing ated for a panel of sample peptides. The COC microfluidic (Su et al. 2005), biomolecular separations performed by chips were fabricated by hot embossing and solvent bond- zone electrophoresis (Chen et al. 2002; Shen et al.
    [Show full text]
  • A Flexible Microfluidic System for Single-Cell Transcriptome Profiling
    www.nature.com/scientificreports OPEN A fexible microfuidic system for single‑cell transcriptome profling elucidates phased transcriptional regulators of cell cycle Karen Davey1,7, Daniel Wong2,7, Filip Konopacki2, Eugene Kwa1, Tony Ly3, Heike Fiegler2 & Christopher R. Sibley 1,4,5,6* Single cell transcriptome profling has emerged as a breakthrough technology for the high‑resolution understanding of complex cellular systems. Here we report a fexible, cost‑efective and user‑ friendly droplet‑based microfuidics system, called the Nadia Instrument, that can allow 3′ mRNA capture of ~ 50,000 single cells or individual nuclei in a single run. The precise pressure‑based system demonstrates highly reproducible droplet size, low doublet rates and high mRNA capture efciencies that compare favorably in the feld. Moreover, when combined with the Nadia Innovate, the system can be transformed into an adaptable setup that enables use of diferent bufers and barcoded bead confgurations to facilitate diverse applications. Finally, by 3′ mRNA profling asynchronous human and mouse cells at diferent phases of the cell cycle, we demonstrate the system’s ability to readily distinguish distinct cell populations and infer underlying transcriptional regulatory networks. Notably this provided supportive evidence for multiple transcription factors that had little or no known link to the cell cycle (e.g. DRAP1, ZKSCAN1 and CEBPZ). In summary, the Nadia platform represents a promising and fexible technology for future transcriptomic studies, and other related applications, at cell resolution. Single cell transcriptome profling has recently emerged as a breakthrough technology for understanding how cellular heterogeneity contributes to complex biological systems. Indeed, cultured cells, microorganisms, biopsies, blood and other tissues can be rapidly profled for quantifcation of gene expression at cell resolution.
    [Show full text]
  • Digital Microfluidics
    Microfluid Nanofluid DOI 10.1007/s10404-007-0161-8 REVIEW Digital microfluidics: is a true lab-on-a-chip possible? R. B. Fair Received: 10 February 2007 / Accepted: 14 February 2007 Ó Springer-Verlag 2007 Abstract The suitability of electrowetting-on-dielectric elemental operations, such as transport, mixing, or dis- (EWD) microfluidics for true lab-on-a-chip applications is pensing. These elemental operations are performed on an discussed. The wide diversity in biomedical applications elemental set of components, such as electrode arrays, can be parsed into manageable components and assembled separation columns, or reservoirs. Examples of the incor- into architecture that requires the advantages of being poration of these principles in complex biomedical appli- programmable, reconfigurable, and reusable. This capa- cations are described. bility opens the possibility of handling all of the protocols that a given laboratory application or a class of applications Keywords Lab-on-a-chip Á Digital microfluidics Á would require. And, it provides a path toward realizing the Biomedical applications Á Detection Á Analysis Á true lab-on-a-chip. However, this capability can only be Electrowetting realized with a complete set of elemental fluidic compo- nents that support all of the required fluidic operations. Architectural choices are described along with the reali- 1 Introduction zation of various biomedical fluidic functions implemented in on-chip electrowetting operations. The current status of Historically, microfluidic devices have been designed from this EWD toolkit is discussed. However, the question re- the bottom up, whereby various fluidic components are mains: which applications can be performed on a digital combined together to achieve a device that performs a microfluidic platform? And, are there other advantages particular application.
    [Show full text]
  • (EWOD): Current Perspectives and Applications in Ensuring Food Safety
    Volume 28 Issue 4 Article 8 2020 Electrowetting-on-dielectric (EWOD): Current perspectives and applications in ensuring food safety Follow this and additional works at: https://www.jfda-online.com/journal Part of the Food Science Commons, Medicinal Chemistry and Pharmaceutics Commons, Pharmacology Commons, and the Toxicology Commons This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 4.0 License. Recommended Citation Barman, Snigdha Roy; Khan, Imran; Chatterjee, Subhodeep; Saha, Subhajit; Choi, Dukhyun; Lee, Sangmin; and Lin, Zong-Hong (2020) "Electrowetting-on-dielectric (EWOD): Current perspectives and applications in ensuring food safety," Journal of Food and Drug Analysis: Vol. 28 : Iss. 4 , Article 8. Available at: https://doi.org/10.38212/2224-6614.1239 This Review Article is brought to you for free and open access by Journal of Food and Drug Analysis. It has been accepted for inclusion in Journal of Food and Drug Analysis by an authorized editor of Journal of Food and Drug Analysis. Electrowetting-on-dielectric (EWOD): Current perspectives and applications in ensuring food safety REVIEW ARTICLE Snigdha Roy Barman a, Imran Khan b, Subhodeep Chatterjee c, Subhajit Saha a, Dukhyun Choi d,**, Sangmin Lee e,***, Zong-Hong Lin a,c,d,* a Institute of Biomedical Engineering, National Tsing Hua University, Hsinchu 30013, Taiwan b Institute of NanoEngineering and Microsystems, National Tsing Hua University, Hsinchu 30013, Taiwan c Department of Power Mechanical Engineering, National Tsing Hua University, Hsinchu 30013, Taiwan d Department of Mechanical Engineering, Kyung Hee University, Yongin, 17104, South Korea e School of Mechanical Engineering, Chung-Ang University, Seoul 06974, South Korea Abstract Digital microfluidic (DMF) platforms have contributed immensely to the development of multifunctional lab-on-chip systems for performing complete sets of biological and analytical assays.
    [Show full text]
  • RNA-Seq and Microfluidic Digital PCR Identification of Transcriptionally Active Spirochetes in Termite Gut Microbial Communities
    4-1 RNA-Seq and microfluidic digital PCR identification of transcriptionally active spirochetes in termite gut microbial communities Abstract CO2-reductive acetogenesis in termite hindguts is a bacterial process with significant impact on the nutrition of wood-feeding termites. Acetogenic spirochetes have been identified as key mediators of acetogenesis. Here, we use high-throughput, short transcript sequencing (RNA-Seq) and microfluidic, multiplex digital PCR to identify uncultured termite gut spirochetes transcribing genes for hydrogenase-linked formate dehydrogenase (FDHH) enzymes, which are required for acetogenic metabolism in the spirochete, Treponema primitia. To assess FDHH gene (fdhF) transcription within the gut community of a wood-feeding termite, we sequenced ca. 28,000,000 short transcript reads of gut microbial community RNA using Illumina Solexa technology. RNA-Seq results indicate that fdhF transcription in the gut is dominated by two fdhF genotypes: ZnD2sec and Zn2cys. This finding was independently corroborated with cDNA inventory and qRT-PCR transcription measurements. We, therefore, propose that RNA-Seq mapping of microbial community transcripts is specific and quantitative. Following transcriptional assessments, we performed microfluidic, multiplex digital PCR on single termite gut bacterial cells to discover the identity of uncultured bacteria encoding fdhF 4-2 genotypes ZnD2sec and Zn2cys. We identified the specific 16S rRNA gene ribotype of the bacterium encoding Zn2cys fdhF and report that the bacterium is a spirochete. Phylogenetic analysis reveals that this uncultured spirochete, like T. primitia, possesses genes for acetogenic metabolism – formyl-tetra-hydrofolate synthetase and both selenocysteine and cysteine variants of formate dehydrogenase. Microfluidic results also imply a spirochetal origin for ZnD2sec fdhF, but further gene pair associations are required for verification.
    [Show full text]