The authors thank Scott Longwell for helpful comments and edits to the manuscript and Robert Puccinelli for device photography. The authors acknowledge generous support from a Beckman Institute Technology Development Grant. K.B. is supported by a NSF GFRP fellowship and the TLI component of the Stanford Clinical and Translational Science Award to Spectrum (NIH TL1 TR 001084); P.F. acknowledges a McCormick and Gabilan Faculty Fellowship.

<ol>
	<li>Duncombe, T. A., Tentori, A. M., Herr, A. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microfluidics:+reframing+biological+enquiry.">Microfluidics: reframing biological enquiry.</a> Nat. Rev. Mol. Cell Bio. 16 (9), (2015).</li><li>Squires, T. M., Quake, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microfluidics:+Fluid+physics+at+the+nanoliter+scale.">Microfluidics: Fluid physics at the nanoliter scale.</a> Rev.Mod. Phys. 77 (3), (2005).</li><li>Whitesides, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+origins+and+the+future+of+microfluidics.">The origins and the future of microfluidics.</a> Nature. 442 (7101), (2006).</li><li>Kalisky, T., Blainey, P., Quake, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genomic+Analysis+at+the+Single-Cell+Level.">Genomic Analysis at the Single-Cell Level.</a> Ann. Rev. of Genetics. 45 (1), (2011).</li><li>Finkel, N. H., Lou, X., Wang, C., He, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Peer+Reviewed:+Barcoding+the+Microworld.">Peer Reviewed: Barcoding the Microworld.</a> Anal. Chem. 76 (19), (2004).</li><li>Lecault, V., White, A. K., Singhal, A., Hansen, C. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microfluidic+single+cell+analysis:+from+promise+to+practice.">Microfluidic single cell analysis: from promise to practice.</a> Curr. Opin. in Chem. Bio. 16 (3-4), (2012).</li><li>White, A. K., Heyries, K. A., Doolin, C., VanInsberghe, M., Hansen, C. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-Throughput+Microfluidic+Single-Cell+Digital+Polymerase+Chain+Reaction.">High-Throughput Microfluidic Single-Cell Digital Polymerase Chain Reaction.</a> Anal. Chem. 85 (15), (2013).</li><li>Hansen, C. L., Classen, S., Berger, J. M., Quake, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Microfluidic+Device+for+Kinetic+Optimization+of+Protein+Crystallization+and+In+Situ+Structure+Determination.">A Microfluidic Device for Kinetic Optimization of Protein Crystallization and In Situ Structure Determination.</a> J. Am. Chem. Soc. 128 (10), (2006).</li><li>Maerkl, S. J., Quake, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Systems+Approach+to+Measuring+the+Binding+Energy+Landscapes+of+Transcription+Factors.">A Systems Approach to Measuring the Binding Energy Landscapes of Transcription Factors.</a> Science. 315 (5809), (2007).</li><li>Fordyce, P. M., Gerber, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=De+novo+identification+and+biophysical+characterization+of+transcription-factor+binding+sites+with+microfluidic+affinity+analysis.">De novo identification and biophysical characterization of transcription-factor binding sites with microfluidic affinity analysis.</a> Nat. Biotech. 28 (9), (2010).</li><li>Fan, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Integrated+barcode+chips+for+rapid,+multiplexed+analysis+of+proteins+in+microliter+quantities+of+blood.">Integrated barcode chips for rapid, multiplexed analysis of proteins in microliter quantities of blood.</a> Nat. Biotech. 26 (12), (2008).</li><li>Kovarik, M. L., Gach, P. C., Ornoff, D. M., Wang, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Micro+total+analysis+systems+for+cell+biology+and+biochemical+assays.">Micro total analysis systems for cell biology and biochemical assays.</a> Anal. Chem. , (2011).</li><li>Stroock, A. D., Dertinger, S. K. W., Ajdari, A., Mezi&#263;, I., Stone, H. A., Whitesides, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chaotic+Mixer+for+Microchannels.">Chaotic Mixer for Microchannels.</a> Science. 295 (5555), 647-651 (2002).</li><li>Unger, M. A., Chou, H. -. P., Thorsen, T., Scherer, A., Quake, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Monolithic+Microfabricated+Valves+and+Pumps+by+Multilayer+Soft+Lithography.">Monolithic Microfabricated Valves and Pumps by Multilayer Soft Lithography.</a> Science. 288 (5463), 113-116 (2000).</li><li>Thorsen, T., Maerkl, S. J., Quake, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microfluidic+Large-Scale+Integration.">Microfluidic Large-Scale Integration.</a> Science. 298 (5593), (2002).</li><li>Li, N., Sip, C., Folch, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microfluidic+Chips+Controlled+with+Elastomeric+Microvalve+Arrays.">Microfluidic Chips Controlled with Elastomeric Microvalve Arrays.</a> JoVE. (8), e296 (2007).</li><li>Kim, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Soft+lithography+for+microfluidics:+a+review.">Soft lithography for microfluidics: a review.</a> Biochip. J. 2 (1), 1-11 (2008).</li><li>Studer, V., Hang, G., Pandolfi, A., Ortiz, M., Anderson, W. F., Quake, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scaling+properties+of+a+low-actuation+pressure+microfluidic+valve.">Scaling properties of a low-actuation pressure microfluidic valve.</a> J. Appl. Phys. 95 (1), 393-398 (2004).</li><li>Kartalov, E. P., Scherer, A., Quake, S. R., Taylor, C. R., Anderson, W. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimentally+validated+quantitative+linear+model+for+the+device+physics+of+elastomeric+microfluidic+valves.">Experimentally validated quantitative linear model for the device physics of elastomeric microfluidic valves.</a> J. Appl. Phys. 101 (6), 064505 (2007).</li><li>Gerver, R. E., G&#243;mez-Sj&#246;berg, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Programmable+microfluidic+synthesis+of+spectrally+encoded+microspheres.">Programmable microfluidic synthesis of spectrally encoded microspheres.</a> Lab. Chip. 12 (22), 4716-4723 (2012).</li><li>Fordyce, P. M., Diaz-Botia, C. A., DeRisi, J. L., G&#243;mez-Sj&#246;berg, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Systematic+characterization+of+feature+dimensions+and+closing+pressures+for+microfluidic+valves+produced+via+photoresist+reflow.">Systematic characterization of feature dimensions and closing pressures for microfluidic valves produced via photoresist reflow.</a> Lab. Chip. 12 (21), 4287-4295 (2012).</li><li>Li, C. -. W., Cheung, C. N., Yang, J., Tzang, C. H., Yang, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=PDMS-based+microfluidic+device+with+multi-height+structures+fabricated+by+single-step+photolithography+using+printed+circuit+board+as+masters.">PDMS-based microfluidic device with multi-height structures fabricated by single-step photolithography using printed circuit board as masters.</a> The Analyst. 128 (9), 1137-1142 (2003).</li><li>Romanowsky, M. B., Abate, A. R., Rotem, A., Holtze, C., Weitz, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High+throughput+production+of+single+core+double+emulsions+in+a+parallelized+microfluidic+device.">High throughput production of single core double emulsions in a parallelized microfluidic device.</a> Lab. Chip. 12 (4), 802-807 (2012).</li><li>Mata, A., Fleischman, A. J., Roy, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fabrication+of+multi-layer+SU-8+microstructures.">Fabrication of multi-layer SU-8 microstructures.</a> JMM. 16 (2), 276 (2006).</li><li>. Rafael's Microfluidics Site Available from: <a target="_blank" href="https://sites.google.com/site/rafaelsmicrofluidicspage/valve-controllers">https://sites.google.com/site/rafaelsmicrofluidicspage/valve-controllers</a> (2016)</li><li>Wanat, S., Plass, R., Sison, E., Zhuang, H., Lu, P. -. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optimized+Thick+Film+Processing+for+Bumping+Layers.">Optimized Thick Film Processing for Bumping Layers.</a> Proc. SPIE. , 1281-1288 (2003).</li></ol>

The authors declare that they have no competing financial interests.

이 작품은 우리의 온라인 도구 (26)와 제조업체 지침 25을 기반으로 제작 매개 변수에 대한 간단한 수정을 모든 응용 프로그램에 맞게 조정 할 수 있습니다 밸브 및 가변 높이 구조와 다층 미세 유체 소자를위한 완벽한 여러 단계의 포토 리소그래피 프로토콜을 보여줍니다. 이 프로토콜은 단순한 수동적 하나의 계층 형 이상 미세 유체 장치를 구성하고자하는 연?...

<html> 지난 15 년 동안, 필드와 같은 마이크로 유체는 마이크로 미터 규모 1 유체의 조작을 가능하게 새로운 기술의 폭발과 함께, 빠른 성장을 겪고있다. 처리량을 급격히 증가 및 스케일 (2, 3)의 경제를 활용하여 비용을 절감하는 동시에 작은 양 증가 속도 및 감도를 실현할 수있는 가능성이 있기 때문에 미세 유체 시스템은 습식 실험실 기능을위한 매력적인 플랫폼이다. 적층 마이크로 유체 시스템들은 단일 세포 분석 4, 5, 6, 단일 분자 분석 (예를 들어, 디지털 PCR 7) 단백질 결정학 8 전사 인자 결합 분석 높은 처리량 생화학 분석 분야에서 특히 중요한 영향을 만들었다F &quot;&gt; 9, 10, 11 및 세포 선별. 마이크로 유체의 중심 목표 총 생화학 분석 (12)는 단일 장치 내에 유체 복잡한 조작을 수행 할 수있는 장치 &quot;칩의 실험실 &#39;을 개발하고있다. 다층 소프트 리소그래피 기술의 발달은 온 - 칩 밸브 믹서의 생성을 가능하게함으로써 이러한 목표를 실현하는 데 도움 적극적 소량 13, 14, (15) 내의 유체를 제어하기위한 펌프 하였다. 자신의 장점과 증명 응용 프로그램에도 불구하고, 이러한 미세 유체 기술의 대부분은 비 전문가 사용자가 크게 unharnessed 남아있다. 광범위한 채택으로 인해 미세 시설에 제한된 액세스 부분에 도전하고 있지만 인해 제조 기술의 부적절한 통신에있다. 이 FO 특히 그러하다연구 밸브 또는 복잡한 형상에 대한 구조를 갖춘 다층 미세 유체 장치 : 중요한 설계 매개 변수 및 제조 기술에 대한 자세한, 실용적인 정보의 소수가 종종 이러한 장치의 설계 및 제작을 포함하는 프로젝트에 착수하기위한 새로운 연구를 방지 할 수 있습니다. 이 문서에서는, 밸브 및 가변 높이 기능 다층 미세 유체 장치를 만드는 설계 매개 변수에서 시작하여 모든 제조 단계를 통해 이동을위한 완벽한 프로토콜을 제시하여이 지식 격차를 해결하는 것을 목표로하고있다. 제조의 초기 포토 리소그래피 단계에 초점을 맞춤으로써,이 프로토콜은 금형에서 장치를 주조 및 특정 실험을 실행하는 다운 스트림 단계를 설명하는 다른 미세 유체 프로토콜 16을 보완한다. 모 놀리 식 온 - 칩 밸브와 마이크로 유체 장치는 두 개의 층으로 구성되어 있습니다 : 관심의 유체가 마이크로에서 조작하는 &quot;흐름&quot;층을,채널, 공기 또는 물을 포함하는 마이크로 채널을 선택적으로 유동 층 (14)의 유체 흐름을 조절할 수있는 &quot;권한&quot;층. 이 두 층은 각각의 후속 호출 과정에서 폴리 디메틸 실록산 (PDMS) 복제 성형에 사용되는 별도의 실리콘 몰딩 마스터 상에 제조되는 &quot;소프트 리소그래피 (17).&quot; 적층 장치를 형성하기 위해 PDMS 층 각각은 각각의 성형 마스터에서 주조 한 다음함으로써 각 층의 채널을 시청할 수있는 복합 PDMS 소자를 형성하고, 서로 정렬. 밸브는 유동 제어 채널이 서로 교차 만 얇은 막에 의해 분리 된 위치에 형성된다; 제어 채널의 가압이 유로를 폐색이 막 편향 로컬 유체 (도 1)를 대체. 액티브 온 - 칩 밸브를 원하는 최종 용도에 따라 여러 가지 방법으로 제조 될 수있다. 밸브제어 층 (도 1) (15) 위 또는 유동 층 아래에 있는지에 따라 &quot;밀어&quot;, &quot;아래로 밀어&quot;또는 구조 중 하나로 구성 될 수있다. 유동 채널 접합 기판과 직접 접촉 할 수 있도록 형상이 허용하는 &quot;푸시 다운&quot;상태에서 기판 표면을 선택적으로 기능화 또는 패터닝의 이점을 부여하는, 기하학 하부 폐쇄 압력과 박리에 대해 더 높은 장치의 안정성을 허용 &quot;밀어&quot; 나중에 기능 18, 19. 밸브는 유로의 단면 형상에 따라 의도적 누설 &quot;시브&quot;밸브 또는 완전히 밀봉 될 수있다. 체 밸브 구슬, 세포 또는 다른 macroanalytes 1을 포착하는 데 유용하며, 전형적인 네거티브 포토 레지스트 (즉, SU-8 시리즈), 하의 사용을 통해 제작된다직사각형 프로필을했습니다. 제어 채널은 이들 밸브 영역 위에 가압시, 제어 흐름 층의 PDMS 막 (그림 1)의 모서리를 밀봉 유체 흐름을 허용하지만, 거시적 입자들을 트래핑하지 않고 밸브의 직사각형 프로파일로 등방성으로 편향시킨다. 반대로, 완전 밀봉 미세 유체 밸브는 밸브 위치에서 둥근 포토 레지스트의 작은 패치를 포함하여 제조된다. 이 구조에 의해, 제어 채널의 완전 가압 유체 흐름을 정지, 채널을 밀봉하기 위하여 둥근 유동 층에 대해 막 편향. 흐름 층에서 둥근 프로파일은 일반적인 포토 리소그래피 단계 후 용융 및 긍정적 인 포토 레지스트 (예를 들어, AZ50의 XT 또는 SPR 220)의 리플 로우를 통해 생성된다. 우리는 이전에 밸브 지역의 후 리플 로우 높이가 선택한 기능 치수 (21)에 달려 있음을 증명하고있다. 이 프로토콜은 두 밸브 형상의 제작과를 보여줍니다비드 합성 장치이다. <img alt="그림 1" src="/files/ftp_upload/55276/55276fig1.jpg" /> 그림 1 : 다층 미세 유체 밸브 기하학. 체에 대한 일반적인 &quot;밀어&quot;장치 아키텍처와 (위)과 후 (아래) 가압 전에 완전히 밀봉 밸브. <a href="http://ecsource.jove.com/files/ftp_upload/55276/55276fig1large.jpg" target="_blank">이 그림의 더 큰 버전을 보려면 여기를 클릭하십시오.</a> 장치는 또한 하나의 흐름 층 내에서 여러 다른 높이의 기능을 필요로 저항 (20) 혼란 믹서 (13)와 온 - 칩 복잡한 수동 기능을 포함 할 수있다. 가변 높이 흐름 층을 달성하기 위해, 다른 그룹은 인쇄 회로 기판 에칭 (22), 다층 PDMS 릴리프 배향 (23) 또는 다단계 (P)을 포함한 여러 방법을 채용 한hotolithography 24. 우리 그룹은 효과적이고 재현 가능한 방법으로 단일 성형 마스터 다단계 포토 리소그래피를 발견했다. 이를 위해, 각 층의 도포 사이에서 발달하지 층에 네가티브 포토 레지스트 (예를 들면, SU-8 포토 레지스트 계)의 두께 채널 구축의 간단한 포토 리소그래피 기술이 이용된다. 각 층은 실리콘 마스터에 두께 사용하여 제조 지침 (25)에 따라 네가티브 포토 레지스트에 방사된다. 이 높이의 특징은 특정 투명 마스크 (도 2) 유리 마스크 판에 부착하고 노광 전의 이전 방사 층 정렬을 사용하여 층상에 패터닝된다. 다단계 포토 리소그래피에있어서, 층간 정밀한 정렬은 전체 높이가 가변 유동 채널을 형성하는데 중요하다. 정렬 후에, 각 층의 두께 의존성 노광 후 베이킹을 실시한다. 현상없이, 다음 층은 SIM이고ilarly 패턴. 이러한 방식으로, 키의 기능은 다수의 마스크의 사용을 통해 단일 유동 웨이퍼 층별을 구축 할 수있다. 각 단계 사이에 개발을 스킵함으로써, 기존의 포토 레지스트 층 (24) (즉, 두 개의 25 ㎛의 층이 50㎛의 기능을 할 수있다) 복합 높이는 기능을 생성하기 위해 사용될 수있다. 또한, 이러한 혼란 믹서 헤링본 홈 (13)의 채널 층의 기능은 이전에 노출 된 기능을 가진 층을 사용하여 제조 될 수있다. 최종 개발 단계는 변수 높이 (그림 3)의 기능을 하나의 흐름 웨이퍼를 만드는 과정을 완료합니다. 여기에서, 온 - 칩 밸브를 복수의 높이와 유체 채널을 제조하는 데 필요한 모든 절차의 예를 포함하는 다단계 포토 리소그래피를위한 완전한 프로토콜이 제공된다. 이 제조 프로토콜 밸브 variab을 필요로하는 다중 층 미세 비드 합성의 문맥에서 제공된다르 높이의 기능을 제공합니다. 이 장치는 Poiseuille 저항 액적 성분을 균질화 혼돈 혼합기 제어를 통해 유량을 조절하기 위해, 오일 시스 내에 온 - 칩 저항기 물방울을 생성하기위한 T-접합부를 포함하며, 모두 완전히 밀봉하고 자체 밸브는 다수의 시약을 포함하는 자동화 된 작업 흐름을 활성화 입력. 다단계 포토 리소그래피를 사용하여, 이러한 기능은 각각의 높이와 포토 레지스트에있어서 상이한 층 상에 제조되며 다음 층이 프로토콜로 구성된다 : (1) 흐름 라운드 밸브 층 (55 μm의, AZ50 XT) (2) 흐름 낮은 층 (55 μm의, SU-8 2,050) (3) 흐름 높은 층 (85 μm의, SU- 8 2025 30 μm의 첨가제 고도), 및 (4) 헤링본 그루브 (125 μm의 2025 SU-8, 40 μm의 첨가제 높이) (도 3). 하이드로 겔 비드 선택적 하류 분석 용 표면 기능화 약물 캡슐화, 라디에이터를 포함한 다양한 용도에 사용될 수있는otracing 및 이미지 분석, 세포 결합; 우리는 이전에 란탄 nanophosphors (20)를 포함하는 분광 엔코딩 PEG 하이드로 겔 비드를 제조하기 위해 이러한 장치의보다 복잡한 버전을 사용했다. 원하는 경우 어떤 실험실 연구 활동에 사용할 수 있도록 여기서 설명하는 디자인은 추가 자료에 포함되어 있습니다. 우리는이 프로토콜은 전문가와 마이크로 유체의 진입 장벽을 낮추고 제조 성공의 기회를 증가하는 밸브 또는 복잡한 형상과 다층 미세 유체 소자 제작에 모두 관심이 비전문가를위한 개방형 자원을 제공 할 것으로 예상된다.

<table border="0" cellpadding="0" cellspacing="0" width="1709">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:459px;">Materials</td>
			<td style="width:332px;"></td>
			<td style="width:259px;"></td>
			<td style="width:659px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Mylar Transparency Masks, 5&#34;</td>
			<td>FineLine Plotting</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">5&#34; Quartz Plates</td>
			<td>United Silica&#160;</td>
			<td>Custom</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">4&#34; Silicon Wafers, Test Grade</td>
			<td>University Wafer</td>
			<td>452</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">SU8 2005, 2025, 2050 photoresist</td>
			<td>Microchem</td>
			<td>Y111045, Y111069, Y111072</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Az50XT&#160;</td>
			<td>Integrated Micromaterials</td>
			<td>AZ50XT-Q</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">SU8 Developer</td>
			<td>Microchem</td>
			<td>Y020100</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">AZ400K 1:3 Developer</td>
			<td>Integrated Micromaterials</td>
			<td>AZ400K1:3-CS</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Pyrex 150 mm glass dish</td>
			<td>Sigma-Aldrich</td>
			<td>CLS3140150-1EA</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Wafer Petri Dishes, 150 mm</td>
			<td>VWR</td>
			<td>25384-326</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Wafer Tweezers&#160;</td>
			<td>Electron Microscopy Sciences (EMS)</td>
			<td>78410-2W</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Trichloro(1H,1H,2H,2H-perfluorooctyl)silane (PFOTS)</td>
			<td>Sigma-Aldrich</td>
			<td>448931-10G</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">2&#34; x 3&#34; glass slides</td>
			<td>Thomas Scientific&#160;</td>
			<td>6686K20</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">RTV 615 elastomeric base and curing agent PDMS set</td>
			<td>Momentive&#160;</td>
			<td>RTV615-1P</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tygon Tubing, 0.02&#34; O.D.&#160;</td>
			<td>Fischer Scientific&#160;</td>
			<td>14-171-284</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Capillary PEEK tubing,&#160; 510 um OD, 125 um ID</td>
			<td>Zeus</td>
			<td>Custom</td>
			<td>360 um PEEK is readily available by Idex (catalog number: 1571)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Cyro 4 mL tube</td>
			<td>Greiner Bio-One</td>
			<td>127279</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Epoxy, 30-minute</td>
			<td>Permatex</td>
			<td>84107</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Metal Pins, 0.025&#34; OD, .013&#34; ID</td>
			<td>New England Small Tube</td>
			<td>NE-1310-02</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Poly(ethylene glycol) diacrylate, Mn 700</td>
			<td>Sigma-Aldrich</td>
			<td>455008-100ML</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Lithium Phenyl(2,4,6-trimethylbenzoyl)phosphinate photoinitator&#160;</td>
			<td>Tokyo Chemical Industry Co.</td>
			<td>L0290</td>
			<td>We typically synthesize LAP in-house.&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">HEPES</td>
			<td>Sigma-Aldrich</td>
			<td>H4034-25G</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Light mineral oil</td>
			<td>Sigma-Aldrich</td>
			<td>330779-1L</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Span-80</td>
			<td>Sigma-Aldrich</td>
			<td>85548</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">ABIL EM 90</td>
			<td>UPI Chem</td>
			<td>420095</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Name&#160;</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:459px;">Equipment</td>
			<td>Equivalent equiptment or homebuilt setups will work equally as well</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Mask Aligner</td>
			<td>Karl Suss</td>
			<td>MA6</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Profilometer</td>
			<td>KLA-Tencor</td>
			<td>Alpha-Step D500</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Spin Coater</td>
			<td>Laurell Technologies</td>
			<td>WS-650-23</td>
			<td>Any spincoater can be used that accepts 100 mm wafers</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Vacuum Dessicator, Bell-Jar Style</td>
			<td>Bel-Art</td>
			<td>420100000</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Oven</td>
			<td>Cole-Palmer</td>
			<td>WU-52120-02</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">UV Spot Curing System with 3 mm LLG option</td>
			<td>Dymax</td>
			<td>41015</td>
			<td>UV LEDs, Xenon Arc Lamps, or other UV sources of the same intensity work equally as well</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">MFCS Microfluidic Fluid Control System</td>
			<td>Fluidgent</td>
			<td>MFCS-EZ</td>
			<td>Syringe pumps, custom pneumatics or other control systems can also be used</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Automated control scripting</td>
			<td>MATLAB</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Hotplate</td>
			<td>Tory Pines Scientific</td>
			<td>HP30</td>
			<td>Any hotplate with uniform heating (i.e. aluminum or ceramic plates) will suffice.&#160;</td>
		</tr>
	</tbody>
</table>

multi-step variable height photolithography for valved multilayer microfluidic devices

Microfluidic systems have enabled powerful new approaches to high-throughput biochemical and biological analysis. However, there remains a barrier to entry for non-specialists who would benefit greatly from the ability to develop their own microfluidic devices to address research questions. Particularly lacking has been the open dissemination of protocols related to photolithography, a key step in the development of a replica mold for the manufacture of polydimethylsiloxane (PDMS) devices. While the fabrication of single height silicon masters has been explored extensively in literature, fabrication steps for more complicated photolithography features necessary for many interesting device functionalities (such as feature rounding to make valve structures, multi-height single-mold patterning, or high aspect ratio definition) are often not explicitly outlined. 
Here, we provide a complete protocol for making multilayer microfluidic devices with valves and complex multi-height geometries, tunable for any application. These fabrication procedures are presented in the context of a microfluidic hydrogel bead synthesizer and demonstrate the production of droplets containing polyethylene glycol (PEG diacrylate) and a photoinitiator that can be polymerized into solid beads. This protocol and accompanying discussion provide a foundation of design principles and fabrication methods that enables development of a wide variety of microfluidic devices. The details included here should allow non-specialists to design and fabricate novel devices, thereby bringing a host of recently developed technologies to their most exciting applications in biological laboratories.

여기서는 액적으로부터 폴리 에틸렌 글리콜 (PEG) 하이드로 겔 비드를 생성 할 수있는 장치를 만들어 판막 가변 높이 미세 다층 형의 제조를 증명 (그림 2). 전체 제조 공정의 개요는도 전작으로부터 설계 요소를 이용하여 (3)에 포함되는 비드의 합성 (1) 층류 변조 (55 μm의) (2) 유동 AZ50의 XT 밸브 둥근 포함한 흐름 층 (4)의 ?...

Watch this Scientific Journal Video about Multi-step Variable Height Photolithography for Valved Multilayer Microfluidic Devices at JoVE.com

Multi-step Variable Height Photolithography for Valved Multilayer Microfluidic Devices

Multilayer microfluidic devices often involve the fabrication of master molds with complex geometries for functionality. This article presents a complete protocol for multi-step photolithography with valves and variable height features tunable to any application. As a demonstration, we fabricate a microfluidic droplet generator capable of producing hydrogel beads.

판막 다층 미세 유체 장치에 대한 여러 단계의 변수 높이 포토 리소그래피

Microfluidic systems have enabled powerful new approaches to high-throughput biochemical and biological analysis. However, there remains a barrier to ...

multi-step-variable-height-photolithography-for-valved-multilayer

Research

JoVE Journal

Bioengineering

14.2K 조회수.  Stanford University. Multilayer microfluidic devices often involve the fabrication of master molds with complex geometries for functionality. This article presents a complete protocol for multi-step photolithography with valves and variable height features tunable to any application. As a demonstration, we fabricate a microfluidic droplet generator capable of producing hydrogel beads. 

비디오: Multi-step Variable Height Photolithography for Valved Multilayer Microfluidic Devices

Separating Beads and Cells in Multi-channel Microfluidic Devices Using Dielectrophoresis and Laminar Flow

Microfluidic devices have advanced cell studies by providing a dynamic fluidic environment on the scale of the cell for studying, manipulating, sorting and counting cells. However, manipulating the cell within the fluidic domain remains a challenge and requires complicated fabrication protocols for forming valves and electrodes, or demands specialty equipment like optical tweezers. Here, we demonstrate that conventional printed circuit boards (PCB) can be used for the non-contact manipulation of cells by employing dielectrophoresis (DEP) for bead and cell manipulation in laminar flow fields for bioactuation, and for cell and bead separation in multichannel microfluidic devices. First, we present the protocol for assembling the DEP electrodes and microfluidic devices, and preparing the cells for DEP. Then, we characterize the DEP operation with polystyrene beads. Lastly, we show representative results of bead and cell separation in a multichannel microfluidic device. In summary, DEP is an effective method for manipulating particles (beads or cells) within microfluidic devices.

Dielectrophoresis (DEP) is an effective method to manipulate cells. Printed circuit boards (PCB) can provide inexpensive, reusable and effective electrodes for contact-free cell manipulation within microfluidic devices. By combining PDMS-based microfluidic channels with coverslips on PCBs, we demonstrate bead and cell manipulation and separation within multichannel microfluidic devices.

멀티 채널 Microfluidic 장치에서 분리 비즈와 세포 Dielectrophoresis와 층류를 사용하여

Microfluidic devices have advanced cell studies by providing a dynamic fluidic environment on the scale of the cell for studying, manipulating, sorting ...

연구

생체공학

Fabrication of Micropatterned Hydrogels for Neural Culture Systems using Dynamic Mask Projection Photolithography

Increasingly, patterned cell culture environments are becoming a relevant technique to study cellular characteristics, and many researchers believe in the need for 3D environments to represent in vitro experiments which better mimic in vivo qualities 1-3. Studies in fields such as cancer research 4, neural engineering 5, cardiac physiology 6, and cell-matrix interaction7,8have shown cell behavior differs substantially between traditional monolayer cultures and 3D constructs.
Hydrogels are used as 3D environments because of their variety, versatility and ability to tailor molecular composition through functionalization 9-12. Numerous techniques exist for creation of constructs as cell-supportive matrices, including electrospinning13, elastomer stamps14, inkjet printing15, additive photopatterning16, static photomask projection-lithography17, and dynamic mask microstereolithography18. Unfortunately, these methods involve multiple production steps and/or equipment not readily adaptable to conventional cell and tissue culture methods. The technique employed in this protocol adapts the latter two methods, using a digital micromirror device (DMD) to create dynamic photomasks for crosslinking geometrically specific poly-(ethylene glycol) (PEG) hydrogels, induced through UV initiated free radical polymerization. The resulting "2.5D" structures provide a constrained 3D environment for neural growth. We employ a dual-hydrogel approach, where PEG serves as a cell-restrictive region supplying structure to an otherwise shapeless but cell-permissive self-assembling gel made from either Puramatrix or agarose. The process is a quick simple one step fabrication which is highly reproducible and easily adapted for use with conventional cell culture methods and substrates.
Whole tissue explants, such as embryonic dorsal root ganglia (DRG), can be incorporated into the dual hydrogel constructs for experimental assays such as neurite outgrowth. Additionally, dissociated cells can be encapsulated in the photocrosslinkable or self polymerizing hydrogel, or selectively adhered to the permeable support membrane using cell-restrictive photopatterning. Using the DMD, we created hydrogel constructs up to ~1mm thick, but thin film (&lt;200 &mu;m) PEG structures were limited by oxygen quenching of the free radical polymerization reaction. We subsequently developed a technique utilizing a layer of oil above the polymerization liquid which allowed thin PEG structure polymerization.
In this protocol, we describe the expeditious creation of 3D hydrogel systems for production of microfabricated neural cell and tissue cultures. The dual hydrogel constructs demonstrated herein represent versatile in vitro models that may prove useful for studies in neuroscience involving cell survival, migration, and/or neurite growth and guidance. Moreover, as the protocol can work for many types of hydrogels and cells, the potential applications are both varied and vast.

Simple techniques are described for the rapid production of microfabricated neural culture systems using a digital micromirror device for dynamic mask projection lithography on regular cell culture substrates. These culture systems may be more representative of natural biological architecture, and the techniques described could be adapted for numerous applications.

동적 마스크 프로젝션 석판술를 사용하여 신경 문화 시스템의 Micropatterned Hydrogels의 제작

Increasingly, patterned cell culture environments are becoming a relevant technique to study cellular characteristics, and many researchers believe in the ...

Simple Microfluidic Devices for in vivo Imaging of C. elegans, Drosophila and Zebrafish

Micro fabricated fluidic devices provide an accessible micro-environment for in vivo studies on small organisms. Simple fabrication processes are available for microfluidic devices using soft lithography techniques 1-3. Microfluidic devices have been used for sub-cellular imaging 4,5, in vivo laser microsurgery 2,6 and cellular imaging 4,7. In vivo imaging requires immobilization of organisms. This has been achieved using suction 5,8, tapered channels 6,7,9, deformable membranes 2-4,10, suction with additional cooling 5, anesthetic gas 11, temperature sensitive gels 12, cyanoacrylate glue 13 and anesthetics such as levamisole 14,15. Commonly used anesthetics influence synaptic transmission 16,17 and are known to have detrimental effects on sub-cellular neuronal transport 4. In this study we demonstrate a membrane based poly-dimethyl-siloxane (PDMS) device that allows anesthetic free immobilization of intact genetic model organisms such as Caenorhabditis elegans (C. elegans), Drosophila larvae and zebrafish larvae. These model organisms are suitable for in vivo studies in microfluidic devices because of their small diameters and optically transparent or translucent bodies. Body diameters range from ~10 &mu;m to ~800 &mu;m for early larval stages of C. elegans and zebrafish larvae and require microfluidic devices of different sizes to achieve complete immobilization for high resolution time-lapse imaging. These organisms are immobilized using pressure applied by compressed nitrogen gas through a liquid column and imaged using an inverted microscope. Animals released from the trap return to normal locomotion within 10 min.
We demonstrate four applications of time-lapse imaging in C. elegans namely, imaging mitochondrial transport in neurons, pre-synaptic vesicle transport in a transport-defective mutant, glutamate receptor transport and Q neuroblast cell division. Data obtained from such movies show that microfluidic immobilization is a useful and accurate means of acquiring in vivo data of cellular and sub-cellular events when compared to anesthetized animals (Figure 1J and 3C-F 4).
Device dimensions were altered to allow time-lapse imaging of different stages of C. elegans, first instar Drosophila larvae and zebrafish larvae. Transport of vesicles marked with synaptotagmin tagged with GFP (syt.eGFP) in sensory neurons shows directed motion of synaptic vesicle markers expressed in cholinergic sensory neurons in intact first instar Drosophila larvae. A similar device has been used to carry out time-lapse imaging of heartbeat in ~30 hr post fertilization (hpf) zebrafish larvae. These data show that the simple devices we have developed can be applied to a variety of model systems to study several cell biological and developmental phenomena in vivo.

Simple Microfluidic Devices for in vivo Imaging of C. elegans, Drosophila and Zebrafish

A simple microfluidic device has been developed to perform anesthetic free in vivo imaging of C. elegans, intact Drosophila larvae and zebrafish larvae. The device utilizes a deformable PDMS membrane to immobilize these model organisms in order to perform time lapse imaging of numerous processes such as heart beat, cell division and sub-cellular neuronal transport. We demonstrate the use of this device and show examples of different types of data collected from different model systems.

에 대한 간단한 마이크로 유체 장치<em&gt; 생체 내</em의&gt; 이미징<em&gt; C. elegans</em&gt;<em&gt; Drosophila</em&gt;와 Zebrafish

Micro fabricated fluidic devices provide an accessible micro-environment for in vivo studies on small organisms. Simple fabrication processes are ...

Multi-step Preparation Technique to Recover Multiple Metabolite Compound Classes for In-depth and Informative Metabolomic Analysis

Metabolomics is an emerging field which enables profiling of samples from living organisms in order to obtain insight into biological processes. A vital aspect of metabolomics is sample preparation whereby inconsistent techniques generate unreliable results. This technique encompasses protein precipitation, liquid-liquid extraction, and solid-phase extraction as a means of fractionating metabolites into four distinct classes.&#160;Improved enrichment of low abundance molecules with a resulting increase in sensitivity is obtained, and ultimately results in more confident identification of molecules. This technique has been applied to plasma, bronchoalveolar lavage fluid, and cerebrospinal fluid samples with volumes as low as 50 &#181;l. &#160;Samples can be used for multiple downstream applications; for example, the pellet resulting from protein precipitation can be stored for later analysis. The supernatant from that step undergoes liquid-liquid extraction using water and strong organic solvent to separate the hydrophilic and hydrophobic compounds. Once fractionated, the hydrophilic layer can be processed for later analysis or discarded if not needed. The hydrophobic fraction is further treated with a series of solvents during three solid-phase extraction steps to separate it into fatty acids, neutral lipids, and phospholipids. This allows the technician the flexibility to choose which class of compounds is preferred for analysis. It also aids in more reliable metabolite identification since some knowledge of chemical class exists.

The reliability of results in metabolomics experiments depends on the effectiveness and reproducibility of the sample preparation. Described is a rigorous and in-depth method that enables extraction of metabolites from biological fluids with the option of subsequently analyzing up to thousands of compounds, or just the compound classes of interest.

심층하고 유익한 대사 체 분석을위한 여러 대사 산물 화합물 클래스를 복구하려면 여러 단계의 제조 기술

Metabolomics is an emerging field which enables profiling of samples from living organisms in order to obtain insight into biological processes. A vital ...

Polymeric Microneedle Array Fabrication by Photolithography

This manuscript describes the fabrication of polymeric microneedle (MN) arrays by photolithography. It involves a simple mold-free process by using a photomask consisting of embedded micro-lenses. Embedded micro-lenses were found to influence MN geometry (sharpness). Robust MN arrays with tip diameters ranging between 41.5 &#181;m &#177; 8.4 &#181;m and 71.6 &#181;m &#177; 13.7 &#181;m, with two different lengths (1,336 &#181;m &#177; 193 &#181;m and 957 &#181;m &#177; 171 &#181;m) were fabricated. These MN arrays may provide potential applications in delivery of low molecular and macromolecular therapeutic agents through skin.

Here, we present a protocol describing a mold-free fabrication process of the polymeric microneedles by photolithography.

포토 리소그래피에 의해 고분자 미세 바늘 어레이 제작

This manuscript describes the fabrication of polymeric microneedle (MN) arrays by photolithography. It involves a simple mold-free process by using a ...

Layer-by-layer Collagen Deposition in Microfluidic Devices for Microtissue Stabilization

Although microfluidics provides exquisite control of the cellular microenvironment, culturing cells within microfluidic devices can be challenging. 3D culture of cells in collagen type I gels helps to stabilize cell morphology and function, which is necessary for creating microfluidic tissue models in microdevices. Translating traditional 3D culture techniques for tissue culture plates to microfluidic devices is often difficult because of the limited channel dimensions. In this method, we describe a technique for modifying native type I collagen to generate polycationic and polyanionic collagen solutions that can be used with layer-by-layer deposition to create ultrathin collagen assemblies on top of cells cultured in microfluidic devices. These thin collagen layers stabilize cell morphology and function, as shown using primary hepatocytes as an example cell, allowing for the long term culture of microtissues in microfluidic devices.

The creation of functional microtissues within microfluidic devices requires the stabilization of cell phenotypes by adapting traditional cell culture techniques to the limited spatial dimensions in microdevices. Modification of collagen allows the layer-by-layer deposition of ultrathin collagen assemblies that can stabilize primary cells, such as hepatocytes, as microfluidic tissue models.

Microtissue 안정화를위한 미세 유체 장치에서 레이어별로 층 콜라겐 증착

Although microfluidics provides exquisite control of the cellular microenvironment, culturing cells within microfluidic devices can be challenging. 3D ...

Using Adhesive Patterning to Construct 3D Paper Microfluidic Devices

We demonstrate the use of patterned aerosol adhesives to construct both planar and nonplanar 3D paper microfluidic devices. By spraying an aerosol adhesive through a metal stencil, the overall amount of adhesive used in assembling paper microfluidic devices can be significantly reduced. We show on a simple 4-layer planar paper microfluidic device that the optimal adhesive application technique and device construction style depends heavily on desired performance characteristics. By moderately increasing the overall area of a device, it is possible to dramatically decrease the wicking time and increase device success rates while also reducing the amount of adhesive required to keep the device together. Such adhesive application also causes the adhesive to form semi-permanent bonds instead of permanent bonds between paper layers, enabling single-use devices to be non-destructively disassembled after use. Nonplanar 3D origami devices also benefit from the semi-permanent bonds during folding, as it reduces the likelihood that unrelated faces may accidently stick together. Like planar devices, nonplanar structures see reduced wicking times with patterned adhesive application vs uniformly applied adhesive.

We demonstrate the use of patterned aerosol adhesives to construct 3D paper microfluidic devices. This method of adhesive application forms semi-permanent bonds between layers, enabling single-use devices to be non-destructively disassembled after use and to ease folding complex nonplanar structures.

3D 종이 미세 유체 장치를 구축하기 위해 접착제 패터닝을 사용하여

We demonstrate the use of patterned aerosol adhesives to construct both planar and nonplanar 3D paper microfluidic devices. By spraying an aerosol ...

Designing Microfluidic Devices for Studying Cellular Responses Under Single or Coexisting Chemical/Electrical/Shear Stress Stimuli

Microfluidic devices are capable of creating a precise and controllable cellular micro-environment of pH, temperature, salt concentration, and other physical or chemical stimuli. They have been commonly used for in vitro cell studies by providing in vivo like surroundings. Especially, how cells response to chemical gradients, electrical fields, and shear stresses has drawn many interests since these phenomena are important in understanding cellular properties and functions. These microfluidic chips can be made of glass substrates, silicon wafers, polydimethylsiloxane (PDMS) polymers, polymethylmethacrylate (PMMA) substrates, or polyethyleneterephthalate (PET) substrates. Out of these materials, PMMA substrates are cheap and can be easily processed using laser ablation and writing. Although a few microfluidic devices have been designed and fabricated for generating multiple, coexisting chemical and electrical stimuli, none of them was considered efficient enough in reducing experimental repeats, particular for screening purposes. In this report, we describe our design and fabrication of two PMMA-based microfluidic chips for investigating cellular responses, in the production of reactive oxygen species and the migration, under single or coexisting chemical/electrical/shear stress stimuli. The first chip generates five relative concentrations of 0, 1/8, 1/2, 7/8, and 1 in the culture regions, together with a shear stress gradient produced inside each of these areas. The second chip generates the same relative concentrations, but with five different electric field strengths created within each culture area. These devices not only provide cells with a precise, controllable micro-environment but also greatly increase the experimental throughput.

Micro-fabricated devices integrated with fluidic components provide an in vitro platform for cell studies mimicking the in vivo micro-environment. We developed polymethylmethacrylate-based microfluidic chips for studying cellular responses under single or coexisting chemical/electrical/shear stress stimuli.

단일 또는 공존 화학 / 전기 / 전단 응력 자극에서 세포 응답 유학을위한 미세 유체 장치 설계

Microfluidic devices are capable of creating a precise and controllable cellular micro-environment of pH, temperature, salt concentration, and other ...

Fabrication of Three-dimensional Paper-based Microfluidic Devices for Immunoassays

Paper wicks fluids autonomously due to capillary action. By patterning paper with hydrophobic barriers, the transport of fluids can be controlled and directed within a layer of paper. Moreover, stacking multiple layers of patterned paper creates sophisticated three-dimensional microfluidic networks that can support the development of analytical and bioanalytical assays. Paper-based microfluidic devices are inexpensive, portable, easy to use, and require no external equipment to operate. As a result, they hold great promise as a platform for point-of-care diagnostics. In order to properly evaluate the utility and analytical performance of paper-based devices, suitable methods must be developed to ensure their manufacture is reproducible and at a scale that is appropriate for laboratory settings. In this manuscript, a method to fabricate a general device architecture that can be used for paper-based immunoassays is described. We use a form of additive manufacturing (multi-layer lamination) to prepare devices that comprise multiple layers of patterned paper and patterned adhesive. In addition to demonstrating the proper use of these three-dimensional paper-based microfluidic devices with an immunoassay for human chorionic gonadotropin (hCG), errors in the manufacturing process that may result in device failures are discussed. We expect this approach to manufacturing paper-based devices will find broad utility in the development of analytical applications designed specifically for limited-resource settings.

We detail a method to fabricate three-dimensional paper-based microfluidic devices for use in the development of immunoassays. Our approach to device assembly is a type of multilayer, additive manufacturing. We demonstrate a sandwich immunoassay to provide representative results for these types of paper-based devices.

면역 측정법에 대한 3 차원 종이 기반 미세 유체 장치의 제작

Paper wicks fluids autonomously due to capillary action. By patterning paper with hydrophobic barriers, the transport of fluids can be controlled and ...

Polydimethylsiloxane-polycarbonate Microfluidic Devices for Cell Migration Studies Under Perpendicular Chemical and Oxygen Gradients

This paper reports a microfluidic device made of polydimethylsiloxane (PDMS) with an embedded polycarbonate (PC) thin film to study cell migration under combinations of chemical and oxygen gradients. Both chemical and oxygen gradients can greatly affect cell migration in vivo; however, due to technical limitations, very little research has been performed to investigate their effects in vitro. The device developed in this research takes advantage of a series of serpentine-shaped channels to generate the desired chemical gradients and exploits a spatially confined chemical reaction method for oxygen gradient generation. The directions of the chemical and oxygen gradients are perpendicular to each other to enable straightforward migration result interpretation. In order to efficiently generate the oxygen gradients with minimal chemical consumption, the embedded PC thin film is utilized as a gas diffusion barrier. The developed microfluidic device can be actuated by syringe pumps and placed into a conventional cell incubator during cell migration experiments to allow for setup simplification and optimized cell culture conditions. In cell experiments, we used the device to study migrations of adenocarcinomic human alveolar basal epithelial cells, A549, under combinations of chemokine (stromal cell-derived factor, SDF-1&#945;) and oxygen gradients. The experimental results show that the device can stably generate perpendicular chemokine and oxygen gradients and is compatible with cells. The migration study results indicate that oxygen gradients may play an essential role in guiding cell migration, and cellular behavior under combinations of gradients cannot be predicted from those under single gradients. The device provides a powerful and practical tool for researchers to study interactions between chemical and oxygen gradients in cell culture, which can promote better cell migration studies in more in vivo-like microenvironments.

The control of chemical and oxygen gradients is essential for cell cultures. This paper reports a polydimethylsiloxane-polycarbonate (PDMS-PC) microfluidic device capable of reliably generating combinations of chemical and oxygen gradients for cell migration studies, which can be practically utilized in biological labs without sophisticated instrumentation.

셀 마이그레이션 연구에서 수직 화학 산소 그라디언트에 대한 폴리 디메틸 실록산 - 폴리 카보네이트 미세 유체 장치

This paper reports a microfluidic device made of polydimethylsiloxane (PDMS) with an embedded polycarbonate (PC) thin film to study cell migration under ...