이 작품은 이스라엘 과학 재단 (부여 번호 1118년에서 1108년까지 및 승인 번호 12분의 1,146)와 미나 크롤 기념 연구 기금에 의해 지원되었다. ES는 기꺼이 러셀 Berrie 나노 기술 연구소의 재정 지원을 인정합니다.

1. 산화 된 다공성 SiO2로 제조 <ol><li> 일정한 전류에서 30 초 동안 수성 HF과 무수 에탄올의 3:1 (V / V) 용액에 에칭 실리콘 웨이퍼 (단 하나 측이 &lt;100&gt;의 얼굴에 광택과 도핑 된 p 형, 0.0008 Ω · cm) 385mA / cm 2의 밀도. HF는 부식성 액체이며, 그것은 극단적 인주의하여 취급해야한다는 것을주의하시기 바랍니다. </li><li> 절대 에탄올을 생성 된 다공성 실리콘 (PSI) 필름의 표면을 여러 번 헹구고, 건조 질소 가스하에 필름을 건조. </li><li> 주변 공기에서 1 시간 동안 800 ° C (해제, 1 시간 동안 노에두고, 800 ° C로 가열로 가열, 실내 온도에서 가열로에서 샘플을 장소에서 튜브로에서 갓 에칭 PSI 샘플을 산화 용광로) 만 실온에서로에서 샘플을 제거합니다. </li></ol> 2. PSiO 2 발판의 Biofunctionalization <ol><li> 산화 된 PSI (PSI를 품어O 2) 메르 캅토에서 1 시간 (톨루엔 트리 메 톡시-3) 95 % (MPTS) 솔루션 (108 MM)에 대한 샘플. </li><li> 톨루엔, 메탄올 및 아세톤으로 실란 처리 PSiO이 샘플을 씻어, 건조 건조 질소 가스하에. </li><li> 100 ㎜ PEO-요오 비오틴 용액 1 ㎖에 30 분간 실란 변성 PSiO이 시료를 배양한다. </li><li> 0.1 M 인산 완충 식염수 (PBS) 용액을 여러 번 비오틴 처리 PSiO 2 샘플을 씻어. </li><li> 100 ㎍ / ㎖의 스트렙 타비 딘 (SA) 용액 1 ㎖에 30 분간 비오틴 - 수정 PSiO 2 샘플을 품어. </li><li> 0.1 M PBS 용액을 여러 번 SA 처리 PSiO 2 샘플을 씻어. </li><li> 100 ㎍ / ㎖의 바이오틴 E. 1 ㎖에 30 분 동안 생성 된 SA-수정 PSiO 2 샘플을 품어 대장균 단일 클론 항체 (면역 글로불린 G, IgG의) 용액 또는 100 ㎍ / ㎖의 바이오틴 - 토끼 IgG를 가진 (단일 클론 항체 모델로). </li></ol> 3. 형광 라벨 및 형광 현미경 <ol><li> 15 ㎍ / ㎖의 형광 (FITC)와 IgG의 수정 표면을 품어 40 분 동안 항 토끼 IgG의 태그 및 15 ㎍ / ㎖의 형광 (FITC)와 컨트롤로 안티 마우스 IgG의 태그. </li><li> 0.1 M PBS 용액을 여러 번와 수정 된 샘플을 씻어. </li><li> 형광 현미경으로 샘플을 검사합니다. </li></ol> 4. 박테리아의 문화 <ol><li> E.에게 경작 대장균 K12의 루리아 베르 타니 (LB) 배지 (: 염화나트륨, 효모 추출물 5 g의 5 g을, 그리고 트립 톤 10 g의 탈 이온수 1 L 중간 구성) 5 ㎖와 10 ㎖ 튜브에 박테리아. 박테리아가 하룻밤 37 ℃에서 진탕 배양한다 </li><li> 600 ㎚의 파장에서의 광학 밀도 (OD)를 판독함으로써 박테리아 농도를 모니터링한다. LB 배지에서 밤새 성장 후, 박테리아의 농도를 결정하는 분광 광도계를 사용하여 OD (600)를 읽는다. 세포의 수는이다 디레OD 600 측정 (1 OD 600 = 10 8 세포 / ml)에 ctly 비례. </li></ol> 5. 박테리아 검출 <ol><li> IgG의 수정 PSiO 2 깔끔한 PSiO 2 (제어)을 맞춤 제작 플렉시 유리 흐름 세포의 샘플을 놓습니다. 샘플 반사율이 모든 측정시 같은 장소에서 측정 될 수 있도록 흐름 ​​셀을 수정합니다. </li><li> E.와 샘플을 품어 실온에서 30 분 동안 대장균 K12 현탁액 (104 세포 / ㎖). 그런 다음 30 분 동안 V 식염수 / W 0.85 %로 셀을 세척하여 세균 현탁액을 제거합니다. </li><li> 실험을하는 동안 반사율 데이터의 변경 사항을 모니터링합니다. 모든 광학 측정은 주변 수계에서 수행 될 필요가있다. 스펙트라는 CCD 분광계를 사용하여 데이터를 수집하고 (25), (26)을 앞서 설명한 바와 같이 고속 푸리에 변환 (FFT)에 변환을 적용함으로써 분석되어야한다. </li><li> 에 박테리아의 존재를 확인할바이오 센서의 표면 바로 바이오 센싱 실험 후 수직 광학 현미경으로 시료의 관찰. </li></ol>

<ol>
	<li>Velusamy, V., 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=An+overview+of+foodborne+pathogen+detection:+In+the+perspective+of+biosensors.">An overview of foodborne pathogen detection: In the perspective of biosensors.</a> Biotechnol. Adv. 28 (2), 232-23 (2010).</li><li>Doyle, M. P., Beuchat, L. R., Montville, T. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Food Microbiol.: Fundamentals and Front. 2, (2001).</li><li>Radke, S. M., Alocilja, E. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+microfabricated+biosensor+for+detecting+foodborne+bioterrorism+agents.">A microfabricated biosensor for detecting foodborne bioterrorism agents.</a> IEEE Sens. J. 5 (4), 744 (2005).</li><li>Glynn, B., 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=Current+and+emerging+molecular+diagnostic+technologies+applicable+to+bacterial+food+safety.">Current and emerging molecular diagnostic technologies applicable to bacterial food safety.</a> Int. J. of Dairy Technol. 59 (2), 126 (2006).</li><li>Leonard, 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=Advances+in+biosensors+for+detection+of+pathogens+in+food+and+water.">Advances in biosensors for detection of pathogens in food and water.</a> Enzyme Microb. Technol. 32 (1), 3 (2003).</li><li>Alvarez, S. 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=Using+a+porous+silicon+photonic+crystal+for+bacterial+cell-based+biosensing.">Using a porous silicon photonic crystal for bacterial cell-based biosensing.</a> Physica Status Solidi a-Applications and Materials Science. 204 (5), 1439 (2007).</li><li>Archer, M., 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=Electrical+porous+silicon+microarray+for+DNA+hybridization+detection.">Electrical porous silicon microarray for DNA hybridization detection.</a> Micro- and Nanosystems. 782, 385 (2004).</li><li>Chan, S., Horner, S. R., Fauchet, P. M., Miller, B. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Identification+of+Gram+Negative+Bacteria+Using+Nanoscale+Silicon+Microcavities.">Identification of Gram Negative Bacteria Using Nanoscale Silicon Microcavities.</a> J. Am. Chem. Soc. 123, 11797 (2001).</li><li>Dancil, K. -. P. S., Greiner, D. P., Sailor, M. J., Canham, L. T., Sailor, M. J., Tanaka, K., Tsai, C. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Development of a Porous Silicon Based Biosensor. 536, 557-562 (1999).</li><li>D'Auria, S., 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=Nanostructured+silicon-based+biosensors+for+the+selective+identification+of+analytes+of+social+interest.">Nanostructured silicon-based biosensors for the selective identification of analytes of social interest.</a> J Phys - Condens Matter. 18 (33), S2019 (2006).</li><li>de Leon, S. B., 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=Neurons+culturing+and+biophotonic+sensing+using+porous+silicon.">Neurons culturing and biophotonic sensing using porous silicon.</a> Appl Phys Lett. 84 (22), 4361 (2004).</li><li>Janshoff, A., 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=Macroporous+p-type+silicon+Fabry-Perot+layers.+Fabrication,+characterization,+and+applications+in+biosensing.">Macroporous p-type silicon Fabry-Perot layers. Fabrication, characterization, and applications in biosensing.</a> J. Am. Chem. Soc. 120 (46), 12108 (1998).</li><li>Orosco, M. M., Pacholski, C., Miskelly, G. M., Sailor, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Protein-coated+porous+silicon+photonic+crystals+for+amplified+optical+detection+of+protease+activity.">Protein-coated porous silicon photonic crystals for amplified optical detection of protease activity.</a> Adv. Mater. 18, 1393 (2006).</li><li>Pacholski, C., 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=Biosensing+using+porous+silicon+double-layer+interferometers:+reflective+interferometric+Fourier+transform+spectroscopy.">Biosensing using porous silicon double-layer interferometers: reflective interferometric Fourier transform spectroscopy.</a> J. Am. Chem. Soc. 127 (33), 11636 (2005).</li><li>Pacholski, C., 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=Reflective+Interferometric+Fourier+Transform+Spectroscopy:+A+Self-Compensating+Label-Free+Immunosensor+Using+Double-layers+of+Porous+SiO2.">Reflective Interferometric Fourier Transform Spectroscopy: A Self-Compensating Label-Free Immunosensor Using Double-layers of Porous SiO2.</a> J. Am. Chem. Soc. 128, 4250 (2006).</li><li>Sailor, M. J., Link, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Smart+Dust:+nanostructured+devices+in+a+grain+of+sand.">Smart Dust: nanostructured devices in a grain of sand.</a> Chem. Comm. , 1375 (2005).</li><li>Schwartz, M. P., Alvarez, S. D., Sailor, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Porous+SiO2+interferometric+biosensor+for+quantitative+determination+of+protein+interactions:+Binding+of+protein+a+to+immunoglobulins+derived+from+different+species.">Porous SiO2 interferometric biosensor for quantitative determination of protein interactions: Binding of protein a to immunoglobulins derived from different species.</a> Anal. Chem. 79 (1), 327 (2007).</li><li>Schwartz, M. i. c. h. a. e. l. 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=The+smart+petri+dish:+A+nanostructured+photonic+crystal+for+real-time+monitoring+of+living+cells.">The smart petri dish: A nanostructured photonic crystal for real-time monitoring of living cells.</a> Langmuir. 22, 7084 (2006).</li><li>Stewart, M. P., Buriak, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chemical+and+biological+applications+of+porous+silicon+technology.">Chemical and biological applications of porous silicon technology.</a> Adv. Mater. 12 (12), 859 (2000).</li><li>Zhang, D., Alocilja, E. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+of+nanoporous+silicon-based+DNA+biosensor+for+the+detection+of+Salmonella+enteritidis.">Characterization of nanoporous silicon-based DNA biosensor for the detection of Salmonella enteritidis.</a> IEEE Sens J. 8 (5-6), 775 (2008).</li><li>Bonanno, L. M., Segal, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nanostructured+porous+silicon-polymer-based+hybrids:+from+biosensing+to+drug+delivery.">Nanostructured porous silicon-polymer-based hybrids: from biosensing to drug delivery.</a> Nanomedicine. 6 (10), 1755 (2011).</li><li>Jane, A., Dronov, R., Hodges, A., Voelcker, N. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Porous+silicon+biosensors+on+the+advance.">Porous silicon biosensors on the advance.</a> Trends Biotechnol. 27 (4), 230 (2009).</li><li>Li, S., Huang, J., Cai, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+porous+silicon+optical+microcavity+for+sensitive+bacteria+detection.">A porous silicon optical microcavity for sensitive bacteria detection.</a> Nanotechnology. 22 (42), 425502 (2011).</li><li>Massad-Ivanir, N., Shtenberg, G., Segal, E., Zahavy, E., Ordentlich, A., Yitzhaki, S., Shafferman, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Nano Bio-Technology for Biomedical and Diagnostics Research. 733, (2012).</li><li>Massad-Ivanir, N., 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=Engineering+Nanostructured+Porous+SiO2+Surfaces+for+Bacteria+Detection+via+"Direct+Cell+Capture".">Engineering Nanostructured Porous SiO2 Surfaces for Bacteria Detection via "Direct Cell Capture".</a> Anal. Chem. 83 (9), 3282-32 (2011).</li><li>Massad-Ivanir, N., Shtenberg, G., Zeidman, T., Segal, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Construction+and+characterization+of+porous+SiO2/hydrogel+hybrids+as+optical+biosensors+for+rapid+detection+of+bacteria.">Construction and characterization of porous SiO2/hydrogel hybrids as optical biosensors for rapid detection of bacteria.</a> Adv Funct Mater. 20 (14), 2269-22 (2010).</li><li>Mathew, F. P., Alocilja, E. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Porous+silicon-based+biosensor+for+pathogen+detection.">Porous silicon-based biosensor for pathogen detection.</a> Biosens. Bioelectron. 20 (8), 1656 (2005).</li><li>Ouyang, H., Archer, M., Fauchet, P. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Frontiers in Surface Nanophotonics. 133, 49 (2007).</li><li>Ouyang, H., DeLouise, L. A., Miller, B. L., Fauchet, P. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Label-free+quantitative+detection+of+protein+using+macroporous+silicon+photonic+bandgap+biosensors.">Label-free quantitative detection of protein using macroporous silicon photonic bandgap biosensors.</a> Anal. Chem. 79 (4), 1502-15 (2007).</li><li>Hermanson, G. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Bioconjugate Techniques. , (1996).</li><li>Piervincenzi, R. T., Reichert, W. M., Hellinga, H. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+engineering+of+a+single-chain+antibody+fragment+for+surface+immobilization+in+an+optical+biosensor.">Genetic engineering of a single-chain antibody fragment for surface immobilization in an optical biosensor.</a> Biosensors and Bioelectronics. 13 (3-4), 305 (1998).</li><li>Saerens, D., Huang, L., Bonroy, K., Muyldermans, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antibody+Fragments+as+Probe+in+Biosensor+Development.">Antibody Fragments as Probe in Biosensor Development.</a> Sensors. 8 (8), 4669 (2008).</li><li>Shtenberg, G., 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=Picking+up+the+Pieces:+A+Generic+Porous+Si+Biosensor+for+Probing+the+Proteolytic+Products+of+Enzymes.">Picking up the Pieces: A Generic Porous Si Biosensor for Probing the Proteolytic Products of Enzymes.</a> Anal. Chem. 85 (3), 1951 (2013).</li><li>Bonanno, L. M., DeLouise, L. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Steric+Crowding+Effects+on+Target+Detection+in+an+Affinity+Biosensor.">Steric Crowding Effects on Target Detection in an Affinity Biosensor.</a> Langmuir. 23 (10), 5817 (2007).</li><li>Banada, P. P., Bhunia, A. K., Mohammed, E., Zourob, S., Turner, A. P. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Principles of Bacterial Detection: Biosensors, Recognition Receptors and Microsystems. , 567 (2008).</li><li>Poma, A., Whitcombe, M., Piletsky, S., Whitcombe, M. J., Piletsky, S. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Designing receptores for the next generation of biosensors. , 105 (2013).</li><li>Dudak, F. C., Boyaci, I. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+an+immunosensor+based+on+surface+plasmon+resonance+for+enumeration+of+Escherichia+coli+in+water+samples.">Development of an immunosensor based on surface plasmon resonance for enumeration of Escherichia coli in water samples.</a> Food Res. Int. 40 (7), 803 (2007).</li><li>Dudak, F. C., Boyaci, I. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+and+label-free+bacteria+detection+by+surface+plasmon+resonance+(SPR)+biosensors.">Rapid and label-free bacteria detection by surface plasmon resonance (SPR) biosensors.</a> Biotechn J. 4 (7), 1003 (2009).</li><li>Skottrup, P. D., Nicolaisen, M., Justesen, A. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Towards+on-site+pathogen+detection+using+antibody-based+sensors.">Towards on-site pathogen detection using antibody-based sensors.</a> Biosens. Bioelectron. 24 (3), 339 (2008).</li><li>Taylor, A. D., Ladd, J., Homola, J., Jiang, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Principles of Bacterial Detection: Biosensors, Recognition Receptors and Microsystems. , 83 (2008).</li></ol>

저자는 더 경쟁 재정적 이익을 선언하지 않습니다.

PSiO 2 나노 구조체 (페롯 박막)에 근거 라벨없는 광 immunosensor가 제작되고, 균 검출을위한 바이오 센서로서의 적용 가능성이 확인된다. 수정 및 문제 해결 immunosensor을 설계의 주요 관심사 중 하나는 바이오 센서 감도 31,32의 감소로 이어질 수 고체 기판 상에 증착 및 패터닝 중에 바람직하지 않은 형태의 변화를 겪는 항체의 감수성이다. 이 ?...

병원성 세균의 초기 정확한 식별은 음식과 물 안전, 환경 모니터링, 점의 배려 진단 1를 위해 매우 중요하다. 전통적인 미생물학 기술은 시간이 힘들고, 소모적이며, &quot;실시간&quot;또는 실험실 환경 외부 미생물을 감지하는 능력을 결여 같이, 바이오 센서는 이러한 과제 2-5을 충족하도록 진화하고있다. 최근 몇 년 동안, 다공성 실리콘 (PSI)은 센서와 바이오 센서 6-20의 디자인을위한 유망한 플랫폼으로 떠오르고있다. 지난 십 년간 PSI 기반의 광 센서 및 바이오 센서에 관한 많은 연구가 (21, 22)를 발표했다. 나노 PSI 층은 전형적으로, 단결정 실리콘 웨이퍼로부터 전기 화학적 양극 에칭에 의해 제조된다. 그 결과 PSI의 나노는 큰 표면 및 무료 볼륨으로 많은 유리한 특징을 전시, 제어 할 수있는 크기와 조정 가능한 OPTI 기공칼 속성 10,16. 이러한 광 발광 8,11과 흰색 빛의 반사율 기반의 간섭 7,19로 PSI 층의 광학적 특성은, 강력하게 환경 조건에 의해 영향을 받는다. 광 발광 스펙트럼 변조 또는 반사율 스펙트럼 (10)의 파장 변화로 관찰 필름의 평균 굴절률 변화에 다공질 층 결과 사이 숙박객 분자 / 분석 대상 물질의 캡처. PSI 광 바이오 센서 기술의 광대 한 혁신 있지만, 박테리아 검출 6,8,20,23-29에 대한 PSI 기반 플랫폼에서만 몇 가지보고가있다. 또, 이러한 개념 증명 연구의 대부분은 &quot;간접&quot;균 검출을 증명하고있다. 따라서, 세포의 용해는 일반적으로 선행 연구 균 29 특성상 표적 단백질 / DNA 단편을 추출하는 것이 요구된다. 우리의 접근 방법은 직접적으로 표적 균을 포착하는 것이다PSI 바이오 센서 상에 세포. 따라서 박테리아를 대상으로 특정 단일 클론 항체는, 다공성 표면에 고정되어있다. 항체 - 항원 상호 작용을 통해, 박테리아 세포의 결합은, 바이오 센서의 표면에 반사율 스펙트럼 24-26의 진폭 (강도)의 변화를 유도한다. 이 작품에서 우리는 광학 PSI 기반의 바이오 센서의 구조에보고하고 대장균의 검출 (대장균) (모델 미생물로 사용) K12 박테리아. 모니터의 레이블이없는 바이오 센서 플랫폼으로의 응용을 보여 광 신호로 인해 페 브리 - 페로 박막 간섭 (그림 1A)에 PSI 나노 구조에서 반사 된 빛이다. 빛의 진폭 / 강도의 변화는 빠른 탐지 및 박테리아의 정량화를 허용, 바이오 센서 표면에 대상 박테리아 세포의 특정 고정 상관 관계가있다.

<table border="1">
	<tbody>
		<tr>
			<td>Name of Reagent/Material</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Si wafer</td>
			<td>Siltronix Corp.</td>
			<td></td>
			<td>Highly-B-doped, p-type, 0.0008 &#937;-cm resistivity, &#60;100&#62; oriented</td>
		</tr>
		<tr>
			<td>Aqueous HF (48%)</td>
			<td>Merck</td>
			<td>101513</td>
			<td></td>
		</tr>
		<tr>
			<td>Ethanol absolute</td>
			<td>Merck</td>
			<td>818760</td>
			<td></td>
		</tr>
		<tr>
			<td>PBS buffer solution (pH 7.4)</td>
			<td></td>
			<td></td>
			<td>prepared by dissolving 50 mM Na2HPO4, 17 mM NaH2PO4, and 68 mM NaCl in Milli-Q water (18.2 M&#937;)</td>
		</tr>
		<tr>
			<td>Saline 0.85% w/v</td>
			<td></td>
			<td></td>
			<td>prepared by dissolving 0.85 g NaCl in 100 ml Milli-Q water (18.2 M&#937;)</td>
		</tr>
		<tr>
			<td>95% (3-Mercaptopropyl)trimethoxysilane (MPTS)</td>
			<td>Sigma Aldrich Chemicals</td>
			<td>175617</td>
			<td></td>
		</tr>
		<tr>
			<td>PEO-iodoacetyl biotin</td>
			<td>Sigma Aldrich Chemicals</td>
			<td>B2059</td>
			<td></td>
		</tr>
		<tr>
			<td>Streptavidin (SA)</td>
			<td>Jackson ImmunoResearch Labs Inc.</td>
			<td>016-000-114</td>
			<td></td>
		</tr>
		<tr>
			<td>Fluorescein (DTAF)-streptavidin</td>
			<td>Jackson ImmunoResearch Labs Inc.</td>
			<td>016-010-084</td>
			<td></td>
		</tr>
		<tr>
			<td>Biotinylated-rabbit IgG</td>
			<td>Jackson ImmunoResearch Labs Inc.</td>
			<td>011-060-003</td>
			<td></td>
		</tr>
		<tr>
			<td>Fluorescently tagged anti-rabbit IgG</td>
			<td>Jackson ImmunoResearch Labs Inc.</td>
			<td>111-095-003</td>
			<td></td>
		</tr>
		<tr>
			<td>Fluorescently tagged anti-mouse IgG</td>
			<td>Jackson ImmunoResearch Labs Inc.</td>
			<td>115-095-003</td>
			<td></td>
		</tr>
		<tr>
			<td>Biotinylated E. coli antibody</td>
			<td>Jackson ImmunoResearch Labs Inc.</td>
			<td>1007</td>
			<td></td>
		</tr>
		<tr>
			<td>E. coli (K-12)</td>
			<td></td>
			<td></td>
			<td>was generously supplied by Prof. Sima Yaron, Technion</td>
		</tr>
	</tbody>
</table>

optical detection of e. coli bacteria by mesoporous silicon biosensors

<html> 나노 다공성 실리콘을 기반으로하는 레이블의 광학 바이오 센서 모델 미생물로, 대장균 K12 박테리아의 빠른 캡처 및 탐지를 위해 설계되었습니다. 연구 시료 (세포 용해에 의해 예)에는 전처리가 필요하지 않습니다 동안 바이오 센서는 표면에 대상 박테리아 세포의 직접 결합에 의존하고있다. 포러스 Si 박막은 바이오 센서의 광 변환 소자로서 사용된다. 백색광 조명 아래에서, 다공성 층은 반사율 스펙트럼에서 잘 해결 페 브리 - 페로 프린지 패턴을 표시합니다. 고속 푸리에 단일 피크의 반사율 데이터 결과로 변환 (FFT)을 적용. FFT 피크의 강도의 변화는 모니터링됩니다. 따라서, 목표 세균은 항체 - 항원 상호 작용을 통해, 바이오 센서의 표면에 포착 박테리아 부속의 &#39;실시간&#39;관측을 허용 FFT 피크의 강도에 측정 가능한 변화를 유발한다. NT &quot;&gt; 전기 화학적 양극 산화 공정에 의해 제조 된 메조 포러스 Si 막는, 목표 세균 특정 단일 클론 항체와 접합된다. 고정화 immunoactivity 및 항체의 특이성은 형광 라벨링 실험에 의해 확인된다. 바이오 센서에 노출되면 타겟 박테리아, 세포를 직접 항체 개질 다공질 실리콘 표면에 포착된다. 이들 특정 캡쳐 이벤트가 바이오 센서의 박막 광학 간섭 스펙트럼의 강도 변화의 원인. 우리는 이러한 바이오 센서 (검출 비교적 낮은 균 농도를 검출 할 수 있음을 입증 시간 미만 10 4 세포 / ml)의 제한입니다.

프로토콜 문자 섹션에서 설명한 산화 PSI (PSiO 2) 필름을 제조한다.도 1b는 열 산화 처리 후의 결과 PSI 필름의 고해상도 주사 전자 현미경 사진을 나타낸다. PSiO 2 층은 30 ~ 80 ㎚의 범위의 직경을 갖는 잘 정의 된 원통형 세공에 의해 특징된다. 단일 클론 항체 (IgG의) 분자는 비오틴-SA 시스템과 함께 잘 설립 실란 화 기술을 사용하여 PSiO 2 표면...

Watch this Scientific Journal Video about Optical Detection of E. coli Bacteria by Mesoporous Silicon Biosensors at JoVE.com

Optical Detection of E. coli Bacteria by Mesoporous Silicon Biosensors

급속한 균 검출 용 라벨없는 광 바이오 센서가 도입된다. 바이오 센서의 표면에 직접 표적 균 세포를 포착하기 위해 디자인 된 나노 다공성 실리콘에 기반하고있다. 우리는 포획 프로브로서, 다공질 변환기에 고정화 된 모노클로 날 항체를 사용한다. 우리의 연구는 (같은 세포 용해와 같은) 더 이전의 샘플 처리와 분 이내에 낮은 세균 농도의 검출에 대해 이러한 바이오 센서의 적용 가능성을 보여줍니다.

의 광학 검출<em&gt; E. 대장균</em메조 포러스 실리콘 바이오 센서에 의해&gt; 박테리아

A label-free optical biosensor based on a nanostructured porous Si is designed for rapid capture and detection of Escherichia coli K12 bacteria, as a ...

optical-detection-of-e-coli-bacteria-by-mesoporous-silicon-biosensors

Research

JoVE Journal

Bioengineering

Optical Detection of E. coli Bacteria by Mesoporous Silicon Biosensors

16.8K 조회수.  Technion - Israel Institute of Technology.  급속한 균 검출 용 라벨없는 광 바이오 센서가 도입된다. 바이오 센서의 표면에 직접 표적 균 세포를 포착하기 위해 디자인 된 나노 다공성 실리콘에 기반하고있다. 우리는 포획 프로브로서, 다공질 변환기에 고정화 된 모노클로 날 항체를 사용한다. 우리의 연구는 (같은 세포 용해와 같은) 더 이전의 샘플 처리와 분 이내에 낮은 세균 농도의 검출에 대해 이러...

비디오: Optical Detection of E. coli Bacteria by Mesoporous Silicon Biosensors

Fabrication of Electrochemical-DNA Biosensors for the Reagentless Detection of Nucleic Acids, Proteins and Small Molecules

As medicine is currently practiced, doctors send specimens to a central laboratory for testing and thus must wait hours or days to 
receive the results. Many patients would be better served by rapid, bedside tests. To this end our laboratory and others have developed a versatile, reagentless 
biosensor platform that supports the quantitative, reagentless, electrochemical detection of nucleic acids (DNA, RNA), proteins (including antibodies) and small 
molecules analytes directly in unprocessed clinical and environmental samples. In this video, we demonstrate the preparation and use of several biosensors in this 
"E-DNA" class. In particular, we fabricate and demonstrate sensors for the detection of a target DNA sequence in a polymerase chain reaction mixture, an HIV-specific antibody and the drug cocaine. The preparation procedure requires only three hours of hands-on effort followed by an overnight incubation, and their use requires only minutes.

"E-DNA" sensors, reagentless, electrochemical biosensors that perform well even when challenged directly in blood and other complex matrices, have been adapted to the detection of a wide range of nucleic acid, protein and small molecule analytes. Here we present a general procedure for the fabrication and use of such sensors.

핵산, 단백질과 작은 분자의 Reagentless 감지를위한 전기 -의 DNA Biosensors의 제작

As medicine is currently practiced, doctors send specimens to a central laboratory for testing and thus must wait hours or days to 
receive the results. ...

연구

생체공학

A Cre-Lox P Recombination Approach for the Detection of Cell Fusion In Vivo

The ability of two or more cells of the same type to fuse has been utilized in metazoans throughout evolution to form many complex organs, including skeletal muscle, bone and placenta. Contemporary studies demonstrate fusion of cells of the same type confers enhanced function. For example, when the trophoblast cells of the placenta fuse to form the syncytiotrophoblast, the syncytiotrophoblast is better able to transport nutrients and hormones across the maternal-fetal barrier than unfused trophoblasts1-4. More recent studies demonstrate fusion of cells of different types can direct cell fate. The "reversion" or modification of cell fate by fusion was once thought to be limited to cell culture systems. But the advent of stem cell transplantation led to the discovery by us and others that stem cells can fuse with somatic cells in vivo and that fusion facilitates stem cell differentiation5-7. Thus, cell fusion is a regulated process capable of promoting cell survival and differentiation and thus could be of central importance for development, repair of tissues and even the pathogenesis of disease.
Limiting the study of cell fusion, is lack of appropriate technology to 1) accurately identify fusion products and to 2) track fusion products over time. Here we present a novel approach to address both limitations via induction of bioluminescence upon fusion (Figure 1); bioluminescence can be detected with high sensitivity in vivo8-15. We utilize a construct encoding the firefly luciferase (Photinus pyralis) gene placed adjacent to a stop codon flanked by LoxP sequences. When cells expressing this gene fuse with cells expressing the Cre recombinase protein, the LoxP sites are cleaved and the stop signal is excised allowing transcription of luciferase. Because the signal is inducible, the incidence of false-positive signals is very low. Unlike existing methods which utilize the Cre/LoxP system16, 17, we have incorporated a "living" detection signal and thereby afford for the first time the opportunity to track the kinetics of cell fusion in vivo.
To demonstrate the approach, mice ubiquitously expressing Cre recombinase served as recipients of stem cells transfected with a construct to express luciferase downstream of a floxed stop codon. Stem cells were transplanted via intramyocardial injection and after transplantation intravital image analysis was conducted to track the presence of fusion products in the heart and surrounding tissues over time. This approach could be adapted to analyze cell fusion in any tissue type at any stage of development, disease or adult tissue repair.

A Cre-Lox P Recombination Approach for the Detection of Cell Fusion In Vivo

A method to track cell fusion in living organisms over time is described. The approach utilizes Cre-LoxP recombination to induce luciferase expression upon cell fusion. The luminescent signal generated can be detected in living organisms using biophotonic imaging systems with a sensitivity of detection of &tilde;1,000 cells in peripheral tissues.

세포 융합의 탐지를위한 Cre - 훈제 연어 P 재조합 접근법 VIVO에서</em

The ability of two or more cells of the same type to fuse has been utilized in metazoans throughout evolution to form many complex organs, including ...

Attaching Biological Probes to Silica Optical Biosensors Using Silane Coupling Agents

In order to interface with biological environments, biosensor platforms, such as the popular Biacore system (based on the Surface Plasmon Resonance (SPR) technique), make use of various surface modification techniques, that can, for example, prevent surface fouling, tune the hydrophobicity / hydrophilicity of the surface, adapt to a variety of electronic environments, and most frequently, induce specificity towards a target of interest.1-5 These techniques extend the functionality of otherwise highly sensitive biosensors to real-world applications in complex environments, such as blood, urine, and wastewater analysis.2,6-7 While commercial biosensing platforms, such as Biacore, have well-understood, standard techniques for performing such surface modifications, these techniques have not been translated in a standardized fashion to other label-free biosensing platforms, such as Whispering Gallery Mode (WGM) optical resonators.8-9
WGM optical resonators represent a promising technology for performing label-free detection of a wide variety of species at ultra-low concentrations.6,10-12 The high sensitivity of these platforms is a result of their unique geometric optics: WGM optical resonators confine circulating light at specific, integral resonance frequencies.13 Like the SPR platforms, the optical field is not totally confined to the sensor device, but evanesces; this "evanescent tail" can then interact with species in the surrounding environment. This interaction causes the effective refractive index of the optical field to change, resulting in a slight, but detectable, shift in the resonance frequency of the device. Because the optical field circulates, it can interact many times with the environment, resulting in an inherent amplification of the signal, and very high sensitivities to minor changes in the environment.2,14-15
To perform targeted detection in complex environments, these platforms must be paired with a probe molecule (usually one half of a binding pair, e.g. antibodies / antigens) through surface modification.2 Although WGM optical resonators can be fabricated in several geometries from a variety of material systems, the silica microsphere is the most common. These microspheres are generally fabricated on the end of an optical fiber, which provides a "stem" by which the microspheres can be handled during functionalization and detection experiments. Silica surface chemistries may be applied to attach probe molecules to their surfaces; however, traditional techniques generated for planar substrates are often not adequate for these three-dimensional structures, as any changes to the surface of the microspheres (dust, contamination, surface defects, and uneven coatings) can have severe, negative consequences on their detection capabilities. Here, we demonstrate a facile approach for the surface functionalization of silica microsphere WGM optical resonators using silane coupling agents to bridge the inorganic surface and the biological environment, by attaching biotin to the silica surface.8,16 Although we use silica microsphere WGM resonators as the sensor system in this report, the protocols are general and can be used to functionalize the surface of any silica device with biotin.

Biosensors interface with complex, biological environments and perform targeted detection by combining highly sensitive sensors with highly specific probes attached to the sensor via surface modification. Here, we demonstrate the surface functionalization of silica optical sensors with biotin using silane coupling agents to bridge the sensor and the biological environment.

실란 커플링 에이전트를 사용하여 실리카 광 Biosensors에 생물 학적 프로브를 부착

In order to interface with biological environments, biosensor platforms, such as the popular Biacore system (based on the Surface Plasmon Resonance (SPR) ...

Analyzing Cellular Internalization of Nanoparticles and Bacteria by Multi-spectral Imaging Flow Cytometry

Nanoparticulate systems have emerged as valuable tools in vaccine delivery through their ability to efficiently deliver cargo, including proteins, to antigen presenting cells1-5. Internalization of nanoparticles (NP) by antigen presenting cells is a critical step in generating an effective immune response to the encapsulated antigen. To determine how changes in nanoparticle formulation impact function, we sought to develop a high throughput, quantitative experimental protocol that was compatible with detecting internalized nanoparticles as well as bacteria. To date, two independent techniques, microscopy and flow cytometry, have been the methods used to study the phagocytosis of nanoparticles. The high throughput nature of flow cytometry generates robust statistical data. However, due to low resolution, it fails to accurately quantify internalized versus cell bound nanoparticles. Microscopy generates images with high spatial resolution; however, it is time consuming and involves small sample sizes6-8. Multi-spectral imaging flow cytometry (MIFC) is a new technology that incorporates aspects of both microscopy and flow cytometry that performs multi-color spectral fluorescence and bright field imaging simultaneously through a laminar core. This capability provides an accurate analysis of fluorescent signal intensities and spatial relationships between different structures and cellular features at high speed.
	Herein, we describe a method utilizing MIFC to characterize the cell populations that have internalized polyanhydride nanoparticles or Salmonella enterica serovar Typhimurium. We also describe the preparation of nanoparticle suspensions, cell labeling, acquisition on an ImageStreamX system and analysis of the data using the IDEAS application. We also demonstrate the application of a technique that can be used to differentiate the internalization pathways for nanoparticles and bacteria by using cytochalasin-D as an inhibitor of actin-mediated phagocytosis.

In this article, we describe a method utilizing multi-spectral imaging flow cytometry to quantify the internalization of polyanhydride nanoparticles or bacteria by RAW 264.7 cells.

멀티 스펙트럼 이미징 유동세포계측법 의해 Nanoparticles과 박테리아의 세포 내면화 분석

Nanoparticulate  systems have emerged as valuable tools in vaccine delivery through their  ability to efficiently deliver cargo, including proteins, to ...

Biosensor for Detection of Antibiotic Resistant Staphylococcus Bacteria

A structurally transformed lytic bacteriophage having a broad host range of Staphylococcus aureus strains and a penicillin-binding protein (PBP 2a) antibody conjugated latex beads have been utilized to create a biosensor designed for discrimination of methicillin resistant (MRSA) and sensitive (MSSA) S. aureus species 1,2. The lytic phages have been converted into phage spheroids by contact with water-chloroform interface. Phage spheroid monolayers have been moved onto a biosensor surface by Langmuir-Blodgett (LB) technique 3. The created biosensors have been examined by a quartz crystal microbalance with dissipation tracking (QCM-D) to evaluate bacteria-phage interactions. Bacteria-spheroid interactions led to reduced resonance frequency and a rise in dissipation energy for both MRSA and MSSA strains. After the bacterial binding, these sensors have been further exposed to the penicillin-binding protein antibody latex beads. Sensors analyzed with MRSA responded to PBP 2a antibody beads; although sensors inspected with MSSA gave no response. This experimental distinction determines an unambiguous discrimination between methicillin resistant and sensitive S. aureus strains. Equally bound and unbound bacteriophages suppress bacterial growth on surfaces and in water suspensions. Once lytic phages are changed into spheroids, they retain their strong lytic activity and show high bacterial capture capability. The phage and phage spheroids can be utilized for testing and sterilization of antibiotic resistant microorganisms. Other applications may include use in bacteriophage therapy and antimicrobial surfaces.

Lytic phage biosensors and antibody beads are able to discriminate between methicillin resistant (MRSA) and sensitive staphylococcus bacteria. The phages were immobilized by a Langmuir-Blodgett method onto a surface of a quartz crystal microbalance sensor and worked as broad range staphylococcus probes. Antibody beads recognize MRSA.

항생제 내성 황색 포도상 구균 박테리아의 검출을위한 바이오 센서

A structurally transformed lytic bacteriophage having a broad host range of Staphylococcus aureus strains and a penicillin-binding protein (PBP 2a) ...

Use of Label-free Optical Biosensors to Detect Modulation of Potassium Channels by G-protein Coupled Receptors

Ion channels control the electrical properties of neurons and other excitable cell types by selectively allowing ions to flow through the plasma membrane1. To regulate neuronal excitability, the biophysical properties of ion channels are modified by signaling proteins and molecules, which often bind to the channels themselves to form a heteromeric channel complex2,3. Traditional assays examining the interaction between channels and regulatory proteins require exogenous labels that can potentially alter the protein&#39;s behavior and decrease the physiological relevance of the target, while providing little information on the time course of interactions in living cells. Optical biosensors, such as the X-BODY Biosciences BIND Scanner system, use a novel label-free technology, resonance wavelength grating (RWG) optical biosensors, to detect changes in resonant reflected light near the biosensor. This assay allows the detection of the relative change in mass within the bottom portion of living cells adherent to the biosensor surface resulting from ligand induced changes in cell adhesion and spreading, toxicity, proliferation, and changes in protein-protein interactions near the plasma membrane. RWG optical biosensors have been used to detect changes in mass near the plasma membrane of cells following activation of G protein-coupled receptors (GPCRs), receptor tyrosine kinases, and other cell surface receptors. Ligand-induced changes in ion channel-protein interactions can also be studied using this assay. In this paper, we will describe the experimental procedure used to detect the modulation of Slack-B sodium-activated potassium (KNa) channels by GPCRs.

Optical biosensor techniques can detect changes in mass near the plasma membrane in living cells and allow one to follow cellular responses in both individual cells and populations of cells. This protocol will describe detection of the modulation of potassium channels by G-protein coupled receptors in intact cells using this approach.

G-단백질 결합 수용체에 의해 칼륨 채널의 변조를 감지하기 위해 라벨이없는 광 바이오 센서의 사용

Ion channels control the electrical properties of neurons and other excitable cell types by selectively allowing ions to flow through the plasma ...

Nanomanipulation of Single RNA Molecules by Optical Tweezers

A large portion of the human genome is transcribed but not translated. In this post genomic era, regulatory functions of RNA have been shown to be increasingly important. As RNA function often depends on its ability to adopt alternative structures, it is difficult to predict RNA three-dimensional structures directly from sequence. Single-molecule approaches show potentials to solve the problem of RNA structural polymorphism by monitoring molecular structures one molecule at a time. This work presents a method to precisely manipulate the folding and structure of single RNA molecules using optical tweezers. First, methods to synthesize molecules suitable for single-molecule mechanical work are described. Next, various calibration procedures to ensure the proper operations of the optical tweezers are discussed. Next, various experiments are explained. To demonstrate the utility of the technique, results of mechanically unfolding RNA hairpins and a single RNA kissing complex are used as evidence. In these examples, the nanomanipulation technique was used to study folding of each structural domain, including secondary and tertiary, independently. Lastly, the limitations and future applications of the method are discussed.

Optical tweezers have been used to study RNA folding by stretching individual molecules from their 5&#8217; and 3&#8217; ends. Here common procedures are described to synthesize RNA molecules for tweezing, calibration of the instrument, and methods to manipulate single molecules.

광 핀셋으로 단일 RNA 분자의 Nanomanipulation

A large portion of the human genome is transcribed but not translated. In this post genomic era, regulatory functions of RNA have been shown to be ...

Label-free Single Molecule Detection Using Microtoroid Optical Resonators

Detecting small concentrations of molecules down to the single molecule limit has impact on areas such as early detection of disease, and fundamental studies on the behavior of molecules. Single molecule detection techniques commonly utilize labels such as fluorescent tags or quantum dots, however, labels are not always available, increase cost and complexity, and can perturb the events being studied. Optical resonators have emerged as a promising means to detect single molecules without the use of labels. Currently the smallest particle detected by a non-plasmonically-enhanced bare optical resonator system in solution is a 25 nm polystyrene sphere1. We have developed a technique known as Frequency Locking Optical Whispering Evanescent Resonator (FLOWER) that can surpass this limit and achieve label-free single molecule detection in aqueous solution2. As signal strength scales with particle volume, our work represents a &#62; 100x improvement in the signal to noise ratio (SNR) over the current state of the art. Here the procedures behind FLOWER are presented in an effort to increase its usage in the field.

We have developed a label-free biosensing system based on optical resonator technology known as Frequency Locking Optical Whispering Evanescent Resonator (FLOWER) that is capable of detecting single molecules in solution. Here the procedures behind this work are described and presented.

Microtoroid 광 공진기를 사용하여 라벨이없는 단일 분자 검출

Detecting small concentrations of molecules down to the single molecule limit has impact on areas such as early detection of disease, and fundamental ...

Multicolor Fluorescence Detection for Droplet Microfluidics Using Optical Fibers

Fluorescence assays are the most common readouts used in droplet microfluidics due to their bright signals and fast time response. Applications such as multiplex assays, enzyme evolution, and molecular biology enhanced cell sorting require the detection of two or more colors of fluorescence. Standard multicolor detection systems that couple free space lasers to epifluorescence microscopes are bulky, expensive, and difficult to maintain. In this paper, we describe a scheme to perform multicolor detection by exciting discrete regions of a microfluidic channel with lasers coupled to optical fibers. Emitted light is collected by an optical fiber coupled to a single photodetector. Because the excitation occurs at different spatial locations, the identity of emitted light can be encoded as a temporal shift, eliminating the need for more complicated light filtering schemes. The system has been used to detect droplet populations containing four unique combinations of dyes and to detect sub-nanomolar concentrations of fluorescein.

Multicolor fluorescence detection in droplet microfluidics typically involves bulky and complex epifluorescence microscope-based detection systems. Here we describe a compact and modular multicolor detection scheme that utilizes an array of optical fibers to temporally encode multicolor data collected by a single photodetector.

물방울 미세 유체에 대한 여러 가지 빛깔의 형광 검출 광학 섬유를 사용하여

Fluorescence assays are the most common readouts used in droplet microfluidics due to their bright signals and fast time response. Applications such as ...

Synthesis of Infectious Bacteriophages in an E. coli-based Cell-free Expression System

A new generation of cell-free transcription-translation (TXTL) systems, engineered to have a greater versatility and modularity, provide novel capabilities to perform basic and applied sciences in test tube reactions. Over the past decade, cell-free TXTL has become a powerful technique for a broad range of novel multidisciplinary research areas related to quantitative and synthetic biology. The new TXTL platforms are particularly useful to construct and interrogate biochemical systems through the execution of synthetic or natural gene circuits. In vitro TXTL has proven convenient to rapidly prototype regulatory elements and biological networks as well as to recapitulate molecular self-assembly mechanisms found in living systems. In this article, we describe how infectious bacteriophages, such as MS2 (RNA), &#934;&#935;174 (ssDNA), and T7 (dsDNA), are entirely synthesized from their genome in one-pot reactions using an all Escherichia coli, cell-free TXTL system. Synthesis of the three coliphages is quantified using the plaque assay. We show how the yield of synthesized phage depends on the biochemical settings of the reactions. Molecular crowding, emulated through a controlled concentration of PEG 8000, affects the amount of synthesized phages by orders of magnitudes. We also describe how to amplify the phages and how to purify their genomes. The set of protocols and results presented in this work should be of interest to multidisciplinary researchers involved in cell-free synthetic biology and bioengineering.

Synthesis of Infectious Bacteriophages in an E. coli-based Cell-free Expression System

A new generation of cell-free transcription-translation platforms has been engineered to construct biochemical systems in vitro through the execution of gene circuits. In this article, we describe how bacteriophages, such as MS2, &#934;&#935;174, and T7, are synthesized from their genome using an all E. coli cell-free TXTL system.