Design and Characterization of Immobilized Enzymes in Microfluidic Systems
Herein we report the fabrication, characterization, and use of total analytical microsystems containing surface-immobilized enzymes. Streptavidin-conjugated alkaline phosphatase was linked to biotinylated phospholipid bilayers coated inside poly(dimethylsiloxane) microchannels and borosilicate micro...
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| Veröffentlicht in: | Analytical chemistry (Washington) 2002-01, Vol.74 (2), p.379-385 |
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| creator | Mao, Hanbin Yang, Tinglu Cremer, Paul S |
| description | Herein we report the fabrication, characterization, and use of total analytical microsystems containing surface-immobilized enzymes. Streptavidin-conjugated alkaline phosphatase was linked to biotinylated phospholipid bilayers coated inside poly(dimethylsiloxane) microchannels and borosilicate microcapillary tubes. Rapid determination of enzyme kinetics at many different substrate concentrations was made possible by carrying out laminar flow-controlled dilution on-chip. This allowed Lineweaver−Burk analysis to be performed from a single experiment with all the data collected simultaneously. The results revealed an enzyme turnover number of 51.1 ± 3.2 s-1 for this heterogeneous system. Furthermore, the same enzyme immobilization strategy was extended to demonstrate that multiple chemical reactions could be performed in sequence by immobilizing various enzymes in series. Specifically, the presence of glucose was detected by two coupled steps employing immobilized avidinD-conjugated glucose oxidase and streptavidin-conjugated horseradish peroxidase. |
| doi_str_mv | 10.1021/ac010822u |
| format | Article |
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Streptavidin-conjugated alkaline phosphatase was linked to biotinylated phospholipid bilayers coated inside poly(dimethylsiloxane) microchannels and borosilicate microcapillary tubes. Rapid determination of enzyme kinetics at many different substrate concentrations was made possible by carrying out laminar flow-controlled dilution on-chip. This allowed Lineweaver−Burk analysis to be performed from a single experiment with all the data collected simultaneously. The results revealed an enzyme turnover number of 51.1 ± 3.2 s-1 for this heterogeneous system. Furthermore, the same enzyme immobilization strategy was extended to demonstrate that multiple chemical reactions could be performed in sequence by immobilizing various enzymes in series. Specifically, the presence of glucose was detected by two coupled steps employing immobilized avidinD-conjugated glucose oxidase and streptavidin-conjugated horseradish peroxidase.</description><identifier>ISSN: 0003-2700</identifier><identifier>EISSN: 1520-6882</identifier><identifier>DOI: 10.1021/ac010822u</identifier><identifier>PMID: 11811412</identifier><identifier>CODEN: ANCHAM</identifier><language>eng</language><publisher>Washington, DC: American Chemical Society</publisher><subject>Bacterial Proteins ; Biological and medical sciences ; Biosensing Techniques - methods ; Enzymes ; Enzymes, Immobilized ; Equipment Design ; Fluids ; Fundamental and applied biological sciences. 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Chem</addtitle><description>Herein we report the fabrication, characterization, and use of total analytical microsystems containing surface-immobilized enzymes. Streptavidin-conjugated alkaline phosphatase was linked to biotinylated phospholipid bilayers coated inside poly(dimethylsiloxane) microchannels and borosilicate microcapillary tubes. Rapid determination of enzyme kinetics at many different substrate concentrations was made possible by carrying out laminar flow-controlled dilution on-chip. This allowed Lineweaver−Burk analysis to be performed from a single experiment with all the data collected simultaneously. The results revealed an enzyme turnover number of 51.1 ± 3.2 s-1 for this heterogeneous system. Furthermore, the same enzyme immobilization strategy was extended to demonstrate that multiple chemical reactions could be performed in sequence by immobilizing various enzymes in series. Specifically, the presence of glucose was detected by two coupled steps employing immobilized avidinD-conjugated glucose oxidase and streptavidin-conjugated horseradish peroxidase.</description><subject>Bacterial Proteins</subject><subject>Biological and medical sciences</subject><subject>Biosensing Techniques - methods</subject><subject>Enzymes</subject><subject>Enzymes, Immobilized</subject><subject>Equipment Design</subject><subject>Fluids</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Glucose</subject><subject>Glucose - analysis</subject><subject>Glucose Oxidase - metabolism</subject><subject>Horseradish Peroxidase</subject><subject>Lipid Bilayers</subject><subject>Microchemistry</subject><subject>Molecular and cellular biology</subject><issn>0003-2700</issn><issn>1520-6882</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2002</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpl0F1rFDEUBuAgFrutXvgHZBAUvBg9J5mZZC7LttsPtih0Be_C2SSjqfNRkxlw99c3sksX7FUu8vBy3pextwifETh-IQMIivPpBZthySGvlOIv2QwARM4lwDE7ifEeABGwesWOERVigXzGbs5d9D_7jHqbzX9RIDO64Lc0-qHPhia77rph7Vu_dTa76LebzsXM99mtN2Fo2slbb7K7TRxdF1-zo4ba6N7s31P2fXGxml_ly6-X1_OzZU5FIcZc1dyWErhCUa_XxhhZAxpuJVqsbWlEIVMXCxWVqKi0tgJqGo6SClcpQeKUfdzlPoThz-TiqDsfjWtb6t0wRS1Ts1qpKsH3_8H7YQp9uk2nOFUhyDqhTzuUCsUYXKMfgu8obDSC_reuflo32Xf7wGndOXuQ-zkT-LAHFA21TaDe-HhwohDpsDK5fOd8Wu7v0z-F37qSQpZ69e1O_zhfLuTlYqVvD7lk4qHE8wMfAeh7mzQ</recordid><startdate>20020115</startdate><enddate>20020115</enddate><creator>Mao, Hanbin</creator><creator>Yang, Tinglu</creator><creator>Cremer, Paul S</creator><general>American Chemical Society</general><scope>BSCLL</scope><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QF</scope><scope>7QO</scope><scope>7QQ</scope><scope>7SC</scope><scope>7SE</scope><scope>7SP</scope><scope>7SR</scope><scope>7TA</scope><scope>7TB</scope><scope>7TM</scope><scope>7U5</scope><scope>7U7</scope><scope>7U9</scope><scope>8BQ</scope><scope>8FD</scope><scope>C1K</scope><scope>F28</scope><scope>FR3</scope><scope>H8D</scope><scope>H8G</scope><scope>H94</scope><scope>JG9</scope><scope>JQ2</scope><scope>KR7</scope><scope>L7M</scope><scope>L~C</scope><scope>L~D</scope><scope>P64</scope><scope>7X8</scope></search><sort><creationdate>20020115</creationdate><title>Design and Characterization of Immobilized Enzymes in Microfluidic Systems</title><author>Mao, Hanbin ; Yang, Tinglu ; Cremer, Paul S</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-a443t-892d57028139bbccc7901c2d71d19d5c347082d06a518a5dd60aff217a4e683a3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2002</creationdate><topic>Bacterial Proteins</topic><topic>Biological and medical sciences</topic><topic>Biosensing Techniques - methods</topic><topic>Enzymes</topic><topic>Enzymes, Immobilized</topic><topic>Equipment Design</topic><topic>Fluids</topic><topic>Fundamental and applied biological sciences. 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| subjects | Bacterial Proteins Biological and medical sciences Biosensing Techniques - methods Enzymes Enzymes, Immobilized Equipment Design Fluids Fundamental and applied biological sciences. Psychology Glucose Glucose - analysis Glucose Oxidase - metabolism Horseradish Peroxidase Lipid Bilayers Microchemistry Molecular and cellular biology |
| title | Design and Characterization of Immobilized Enzymes in Microfluidic Systems |
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