A thiol-ene microfluidic device enabling continuous enzymatic digestion and electrophoretic separation as front-end to mass spectrometric peptide analysis
One of the most attractive aspects of microfluidic chips is their capability of integrating several functional units into one single platform. In particular, enzymatic digestion and chemical separation are important steps in processing samples for many biochemical assays. This study presents the dev...
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| Veröffentlicht in: | Analytical and bioanalytical chemistry 2020-06, Vol.412 (15), p.3559-3571 |
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| creator | Lu, Nan Sticker, Drago Kretschmann, Andreas Petersen, Nickolaj J. Kutter, Jörg P. |
| description | One of the most attractive aspects of microfluidic chips is their capability of integrating several functional units into one single platform. In particular, enzymatic digestion and chemical separation are important steps in processing samples for many biochemical assays. This study presents the development and application of a free-flow electrophoresis microfluidic chip, and its upstream combination with an enzyme microreactor with immobilized pepsin in the same miniaturized platform. The whole microfluidic chip was fabricated by making use of thiol-ene click chemistry. As a proof of concept, different fluorescent dyes and labeled amino acids were continuously separated in the 2D electrophoretic channel. The protease pepsin was immobilized using a covalent linkage with ascorbic acid onto a high-surface monolithic support, also made of thiol-ene. To show the potential of the microfluidic chip for continuous sample preparation and analysis, an oligopeptide was enzymatically digested, and the resulting fragments were separated and collected in a single step (prior to mass spectrometric detection), without the need of further time-consuming liquid handling steps. |
| doi_str_mv | 10.1007/s00216-020-02609-5 |
| format | Article |
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In particular, enzymatic digestion and chemical separation are important steps in processing samples for many biochemical assays. This study presents the development and application of a free-flow electrophoresis microfluidic chip, and its upstream combination with an enzyme microreactor with immobilized pepsin in the same miniaturized platform. The whole microfluidic chip was fabricated by making use of thiol-ene click chemistry. As a proof of concept, different fluorescent dyes and labeled amino acids were continuously separated in the 2D electrophoretic channel. The protease pepsin was immobilized using a covalent linkage with ascorbic acid onto a high-surface monolithic support, also made of thiol-ene. To show the potential of the microfluidic chip for continuous sample preparation and analysis, an oligopeptide was enzymatically digested, and the resulting fragments were separated and collected in a single step (prior to mass spectrometric detection), without the need of further time-consuming liquid handling steps.</description><identifier>ISSN: 1618-2642</identifier><identifier>EISSN: 1618-2650</identifier><identifier>DOI: 10.1007/s00216-020-02609-5</identifier><identifier>PMID: 32253474</identifier><language>eng</language><publisher>Berlin/Heidelberg: Springer Berlin Heidelberg</publisher><subject>Amino acids ; Analysis ; Analytical Chemistry ; Animals ; Ascorbic acid ; Biochemistry ; Characterization and Evaluation of Materials ; Chemical separation ; Chemical synthesis ; Chemistry ; Chemistry and Materials Science ; Click Chemistry ; Digestion ; Electrophoresis ; Electrophoresis, Microchip - instrumentation ; Enzymes ; Enzymes, Immobilized - chemistry ; Equipment Design ; Fluorescent dyes ; Fluorescent indicators ; Food Science ; Lab-On-A-Chip Devices ; Laboratory Medicine ; Mass spectrometry ; Mass Spectrometry - instrumentation ; Microfluidic devices ; Microfluidics ; Monitoring/Environmental Analysis ; Paper in Forefront ; Pepsin ; Pepsin A - chemistry ; Peptides ; Peptides - analysis ; Peptides - isolation & purification ; Proteolysis ; Sample preparation ; Spectrometry ; Sulfhydryl Compounds - chemistry ; Swine ; Thiols</subject><ispartof>Analytical and bioanalytical chemistry, 2020-06, Vol.412 (15), p.3559-3571</ispartof><rights>Springer-Verlag GmbH Germany, part of Springer Nature 2020</rights><rights>COPYRIGHT 2020 Springer</rights><rights>Springer-Verlag GmbH Germany, part of Springer Nature 2020.</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c479t-4f3840eed43a254588c0d71675812f149681ee907a3dfd2c94c6b7835ace108e3</citedby><cites>FETCH-LOGICAL-c479t-4f3840eed43a254588c0d71675812f149681ee907a3dfd2c94c6b7835ace108e3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://link.springer.com/content/pdf/10.1007/s00216-020-02609-5$$EPDF$$P50$$Gspringer$$H</linktopdf><linktohtml>$$Uhttps://link.springer.com/10.1007/s00216-020-02609-5$$EHTML$$P50$$Gspringer$$H</linktohtml><link.rule.ids>314,780,784,27923,27924,41487,42556,51318</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/32253474$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Lu, Nan</creatorcontrib><creatorcontrib>Sticker, Drago</creatorcontrib><creatorcontrib>Kretschmann, Andreas</creatorcontrib><creatorcontrib>Petersen, Nickolaj J.</creatorcontrib><creatorcontrib>Kutter, Jörg P.</creatorcontrib><title>A thiol-ene microfluidic device enabling continuous enzymatic digestion and electrophoretic separation as front-end to mass spectrometric peptide analysis</title><title>Analytical and bioanalytical chemistry</title><addtitle>Anal Bioanal Chem</addtitle><addtitle>Anal Bioanal Chem</addtitle><description>One of the most attractive aspects of microfluidic chips is their capability of integrating several functional units into one single platform. In particular, enzymatic digestion and chemical separation are important steps in processing samples for many biochemical assays. This study presents the development and application of a free-flow electrophoresis microfluidic chip, and its upstream combination with an enzyme microreactor with immobilized pepsin in the same miniaturized platform. The whole microfluidic chip was fabricated by making use of thiol-ene click chemistry. As a proof of concept, different fluorescent dyes and labeled amino acids were continuously separated in the 2D electrophoretic channel. The protease pepsin was immobilized using a covalent linkage with ascorbic acid onto a high-surface monolithic support, also made of thiol-ene. To show the potential of the microfluidic chip for continuous sample preparation and analysis, an oligopeptide was enzymatically digested, and the resulting fragments were separated and collected in a single step (prior to mass spectrometric detection), without the need of further time-consuming liquid handling steps.</description><subject>Amino acids</subject><subject>Analysis</subject><subject>Analytical Chemistry</subject><subject>Animals</subject><subject>Ascorbic acid</subject><subject>Biochemistry</subject><subject>Characterization and Evaluation of Materials</subject><subject>Chemical separation</subject><subject>Chemical synthesis</subject><subject>Chemistry</subject><subject>Chemistry and Materials Science</subject><subject>Click Chemistry</subject><subject>Digestion</subject><subject>Electrophoresis</subject><subject>Electrophoresis, Microchip - instrumentation</subject><subject>Enzymes</subject><subject>Enzymes, Immobilized - chemistry</subject><subject>Equipment Design</subject><subject>Fluorescent dyes</subject><subject>Fluorescent indicators</subject><subject>Food Science</subject><subject>Lab-On-A-Chip Devices</subject><subject>Laboratory Medicine</subject><subject>Mass spectrometry</subject><subject>Mass Spectrometry - instrumentation</subject><subject>Microfluidic devices</subject><subject>Microfluidics</subject><subject>Monitoring/Environmental Analysis</subject><subject>Paper in Forefront</subject><subject>Pepsin</subject><subject>Pepsin A - chemistry</subject><subject>Peptides</subject><subject>Peptides - analysis</subject><subject>Peptides - isolation & purification</subject><subject>Proteolysis</subject><subject>Sample preparation</subject><subject>Spectrometry</subject><subject>Sulfhydryl Compounds - chemistry</subject><subject>Swine</subject><subject>Thiols</subject><issn>1618-2642</issn><issn>1618-2650</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2020</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><recordid>eNp9ktFqFTEQhhdRbK2-gBcS8MabrUk22c1eHoq2QsEbvQ45yexpym6yJlnh-Cg-rbPd2qKIhJCQ-ebPDPNX1WtGzxml3ftMKWdtTTnF3dK-lk-qU9YyVfNW0qcPd8FPqhc531LKpGLt8-qk4Vw2ohOn1c8dKTc-jjUEIJO3KQ7j4p23xMF3b4FAMPvRhwOxMRQflrhkfPtxnExZIX-AXHwMxARHYARbUpxvYoI1mmE2yWzhTIaECviPIyWSyeRM8nzHT1AS0jPMxTtAJTMes88vq2eDGTO8uj_Pqq8fP3y5uKqvP19-uthd11Z0fanF0ChBAZxoDJdCKmWp61jbYa98YKJvFQPoaWcaNzhue2HbfacaaSwwqqA5q95tunOK3xZsR08-WxhHEwC71bxRHZdSSYHo27_Q27gkrBcpQTkXPRfykTqYEbQPQyzJ2FVU71pU4x1OAanzf1C4HOAcYoDB4_sfCXxLwCHlnGDQc_KTSUfNqF4NoTdDaDSEvjOEXmt5c1_xsp_APaT8dgACzQZkDIUDpMeW_iP7CwCowhU</recordid><startdate>20200601</startdate><enddate>20200601</enddate><creator>Lu, Nan</creator><creator>Sticker, Drago</creator><creator>Kretschmann, Andreas</creator><creator>Petersen, Nickolaj J.</creator><creator>Kutter, Jörg P.</creator><general>Springer Berlin Heidelberg</general><general>Springer</general><general>Springer Nature B.V</general><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>3V.</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>7U5</scope><scope>7U7</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8BQ</scope><scope>8FD</scope><scope>8FE</scope><scope>8FG</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABJCF</scope><scope>ABUWG</scope><scope>AEUYN</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BGLVJ</scope><scope>BHPHI</scope><scope>C1K</scope><scope>CCPQU</scope><scope>D1I</scope><scope>DWQXO</scope><scope>F28</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>H8D</scope><scope>H8G</scope><scope>HCIFZ</scope><scope>JG9</scope><scope>JQ2</scope><scope>K9.</scope><scope>KB.</scope><scope>KR7</scope><scope>L7M</scope><scope>LK8</scope><scope>L~C</scope><scope>L~D</scope><scope>M0S</scope><scope>M1P</scope><scope>M7P</scope><scope>P64</scope><scope>PDBOC</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>7X8</scope></search><sort><creationdate>20200601</creationdate><title>A thiol-ene microfluidic device enabling continuous enzymatic digestion and electrophoretic separation as front-end to mass spectrometric peptide analysis</title><author>Lu, Nan ; Sticker, Drago ; Kretschmann, Andreas ; Petersen, Nickolaj J. ; Kutter, Jörg P.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c479t-4f3840eed43a254588c0d71675812f149681ee907a3dfd2c94c6b7835ace108e3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2020</creationdate><topic>Amino acids</topic><topic>Analysis</topic><topic>Analytical Chemistry</topic><topic>Animals</topic><topic>Ascorbic acid</topic><topic>Biochemistry</topic><topic>Characterization and Evaluation of Materials</topic><topic>Chemical separation</topic><topic>Chemical synthesis</topic><topic>Chemistry</topic><topic>Chemistry and Materials Science</topic><topic>Click Chemistry</topic><topic>Digestion</topic><topic>Electrophoresis</topic><topic>Electrophoresis, Microchip - 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Academic</collection><jtitle>Analytical and bioanalytical chemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Lu, Nan</au><au>Sticker, Drago</au><au>Kretschmann, Andreas</au><au>Petersen, Nickolaj J.</au><au>Kutter, Jörg P.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>A thiol-ene microfluidic device enabling continuous enzymatic digestion and electrophoretic separation as front-end to mass spectrometric peptide analysis</atitle><jtitle>Analytical and bioanalytical chemistry</jtitle><stitle>Anal Bioanal Chem</stitle><addtitle>Anal Bioanal Chem</addtitle><date>2020-06-01</date><risdate>2020</risdate><volume>412</volume><issue>15</issue><spage>3559</spage><epage>3571</epage><pages>3559-3571</pages><issn>1618-2642</issn><eissn>1618-2650</eissn><abstract>One of the most attractive aspects of microfluidic chips is their capability of integrating several functional units into one single platform. In particular, enzymatic digestion and chemical separation are important steps in processing samples for many biochemical assays. This study presents the development and application of a free-flow electrophoresis microfluidic chip, and its upstream combination with an enzyme microreactor with immobilized pepsin in the same miniaturized platform. The whole microfluidic chip was fabricated by making use of thiol-ene click chemistry. As a proof of concept, different fluorescent dyes and labeled amino acids were continuously separated in the 2D electrophoretic channel. The protease pepsin was immobilized using a covalent linkage with ascorbic acid onto a high-surface monolithic support, also made of thiol-ene. To show the potential of the microfluidic chip for continuous sample preparation and analysis, an oligopeptide was enzymatically digested, and the resulting fragments were separated and collected in a single step (prior to mass spectrometric detection), without the need of further time-consuming liquid handling steps.</abstract><cop>Berlin/Heidelberg</cop><pub>Springer Berlin Heidelberg</pub><pmid>32253474</pmid><doi>10.1007/s00216-020-02609-5</doi><tpages>13</tpages></addata></record> |
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| subjects | Amino acids Analysis Analytical Chemistry Animals Ascorbic acid Biochemistry Characterization and Evaluation of Materials Chemical separation Chemical synthesis Chemistry Chemistry and Materials Science Click Chemistry Digestion Electrophoresis Electrophoresis, Microchip - instrumentation Enzymes Enzymes, Immobilized - chemistry Equipment Design Fluorescent dyes Fluorescent indicators Food Science Lab-On-A-Chip Devices Laboratory Medicine Mass spectrometry Mass Spectrometry - instrumentation Microfluidic devices Microfluidics Monitoring/Environmental Analysis Paper in Forefront Pepsin Pepsin A - chemistry Peptides Peptides - analysis Peptides - isolation & purification Proteolysis Sample preparation Spectrometry Sulfhydryl Compounds - chemistry Swine Thiols |
| title | A thiol-ene microfluidic device enabling continuous enzymatic digestion and electrophoretic separation as front-end to mass spectrometric peptide analysis |
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