Double-Tagging Polymerase Chain Reaction with a Thiolated Primer and Electrochemical Genosensing based on Gold Nanocomposite Sensor for Food Safety
A novel material for electrochemical biosensing based on rigid conducting gold nanocomposite (nano-AuGEC) is presented. Islands of chemisorbing material (gold nanoparticles) surrounded by nonreactive, rigid, and conducting graphite epoxy composite are thus achieved to avoid the stringent control of...
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| Veröffentlicht in: | Analytical chemistry (Washington) 2009-02, Vol.81 (4), p.1332-1339 |
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| creator | Brasil de Oliveira Marques, Paulo R Lermo, Anabel Campoy, Susana Yamanaka, Hideko Barbé, Jordi Alegret, Salvador Pividori, M. Isabel |
| description | A novel material for electrochemical biosensing based on rigid conducting gold nanocomposite (nano-AuGEC) is presented. Islands of chemisorbing material (gold nanoparticles) surrounded by nonreactive, rigid, and conducting graphite epoxy composite are thus achieved to avoid the stringent control of surface coverage parameters required during immobilization of thiolated oligos in continuous gold surfaces. The spatial resolution of the immobilized thiolated DNA was easily controlled by merely varying the percentage of gold nanoparticles in the composition of the composite. As low as 9 fmol (60 pM) of synthetic DNA were detected in hybridization experiments when using a thiolated probe. Moreover, for the first time a double tagging PCR strategy was performed with a thiolated primer for the detection of Salmonella sp., one of the most important foodborne pathogens affecting food safety. This assay was performed by double-labeling the amplicon during the PCR with a −DIG and −SH set of labeled primers. The thiolated end allows the immobilization of the amplicon on the nano-AuGEC electrode, while digoxigenin allows the electrochemical detection with the antiDIG-HRP reporter in the femtomole range. Rigid conducting gold nanocomposite represents a good material for the improved and oriented immobilization of biomolecules with excellent transducing properties for the construction of a wide range of electrochemical biosensors such as immunosensors, genosensors, and enzymosensors. |
| doi_str_mv | 10.1021/ac801736b |
| format | Article |
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Isabel</creator><creatorcontrib>Brasil de Oliveira Marques, Paulo R ; Lermo, Anabel ; Campoy, Susana ; Yamanaka, Hideko ; Barbé, Jordi ; Alegret, Salvador ; Pividori, M. Isabel</creatorcontrib><description>A novel material for electrochemical biosensing based on rigid conducting gold nanocomposite (nano-AuGEC) is presented. Islands of chemisorbing material (gold nanoparticles) surrounded by nonreactive, rigid, and conducting graphite epoxy composite are thus achieved to avoid the stringent control of surface coverage parameters required during immobilization of thiolated oligos in continuous gold surfaces. The spatial resolution of the immobilized thiolated DNA was easily controlled by merely varying the percentage of gold nanoparticles in the composition of the composite. As low as 9 fmol (60 pM) of synthetic DNA were detected in hybridization experiments when using a thiolated probe. Moreover, for the first time a double tagging PCR strategy was performed with a thiolated primer for the detection of Salmonella sp., one of the most important foodborne pathogens affecting food safety. This assay was performed by double-labeling the amplicon during the PCR with a −DIG and −SH set of labeled primers. The thiolated end allows the immobilization of the amplicon on the nano-AuGEC electrode, while digoxigenin allows the electrochemical detection with the antiDIG-HRP reporter in the femtomole range. Rigid conducting gold nanocomposite represents a good material for the improved and oriented immobilization of biomolecules with excellent transducing properties for the construction of a wide range of electrochemical biosensors such as immunosensors, genosensors, and enzymosensors.</description><identifier>ISSN: 0003-2700</identifier><identifier>EISSN: 1520-6882</identifier><identifier>DOI: 10.1021/ac801736b</identifier><identifier>PMID: 19170528</identifier><identifier>CODEN: ANCHAM</identifier><language>eng</language><publisher>Washington, DC: American Chemical Society</publisher><subject>Analytical chemistry ; Animals ; Base Sequence ; Biological and medical sciences ; Biosensing Techniques - methods ; Biosensors ; Biotechnology ; Chemistry ; Deoxyribonucleic acid ; Digoxigenin - chemistry ; Digoxigenin - metabolism ; DNA ; DNA Primers - genetics ; DNA Primers - metabolism ; DNA, Bacterial - chemistry ; DNA, Bacterial - genetics ; DNA, Bacterial - metabolism ; Electrochemical methods ; Electrochemistry ; Electrodes ; Exact sciences and technology ; Food Analysis - methods ; Food Microbiology ; Food safety ; Fundamental and applied biological sciences. Psychology ; Genetic engineering ; Genetic technics ; Genome, Bacterial ; Gold ; Gold - chemistry ; In vitro gene amplification. Pcr technique ; Methods. Procedures. Technologies ; Nanocomposites ; Nanocomposites - chemistry ; Nucleic Acid Hybridization ; Polymerase Chain Reaction ; Salmonella ; Salmonella - genetics ; Salmonella - isolation & purification ; Sulfhydryl Compounds - chemistry ; Surface Properties ; Time Factors ; Various methods and equipments</subject><ispartof>Analytical chemistry (Washington), 2009-02, Vol.81 (4), p.1332-1339</ispartof><rights>Copyright © 2009 American Chemical Society</rights><rights>2009 INIST-CNRS</rights><rights>Copyright American Chemical Society Feb 15, 2009</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-a370t-d8f11acf05733d649407d8b8740bf022b2e99ec96a0e642f46e526ffa26d2383</citedby><cites>FETCH-LOGICAL-a370t-d8f11acf05733d649407d8b8740bf022b2e99ec96a0e642f46e526ffa26d2383</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://pubs.acs.org/doi/pdf/10.1021/ac801736b$$EPDF$$P50$$Gacs$$H</linktopdf><linktohtml>$$Uhttps://pubs.acs.org/doi/10.1021/ac801736b$$EHTML$$P50$$Gacs$$H</linktohtml><link.rule.ids>314,780,784,2765,27076,27924,27925,56738,56788</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=21174116$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/19170528$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Brasil de Oliveira Marques, Paulo R</creatorcontrib><creatorcontrib>Lermo, Anabel</creatorcontrib><creatorcontrib>Campoy, Susana</creatorcontrib><creatorcontrib>Yamanaka, Hideko</creatorcontrib><creatorcontrib>Barbé, Jordi</creatorcontrib><creatorcontrib>Alegret, Salvador</creatorcontrib><creatorcontrib>Pividori, M. Isabel</creatorcontrib><title>Double-Tagging Polymerase Chain Reaction with a Thiolated Primer and Electrochemical Genosensing based on Gold Nanocomposite Sensor for Food Safety</title><title>Analytical chemistry (Washington)</title><addtitle>Anal. Chem</addtitle><description>A novel material for electrochemical biosensing based on rigid conducting gold nanocomposite (nano-AuGEC) is presented. Islands of chemisorbing material (gold nanoparticles) surrounded by nonreactive, rigid, and conducting graphite epoxy composite are thus achieved to avoid the stringent control of surface coverage parameters required during immobilization of thiolated oligos in continuous gold surfaces. The spatial resolution of the immobilized thiolated DNA was easily controlled by merely varying the percentage of gold nanoparticles in the composition of the composite. As low as 9 fmol (60 pM) of synthetic DNA were detected in hybridization experiments when using a thiolated probe. Moreover, for the first time a double tagging PCR strategy was performed with a thiolated primer for the detection of Salmonella sp., one of the most important foodborne pathogens affecting food safety. This assay was performed by double-labeling the amplicon during the PCR with a −DIG and −SH set of labeled primers. The thiolated end allows the immobilization of the amplicon on the nano-AuGEC electrode, while digoxigenin allows the electrochemical detection with the antiDIG-HRP reporter in the femtomole range. Rigid conducting gold nanocomposite represents a good material for the improved and oriented immobilization of biomolecules with excellent transducing properties for the construction of a wide range of electrochemical biosensors such as immunosensors, genosensors, and enzymosensors.</description><subject>Analytical chemistry</subject><subject>Animals</subject><subject>Base Sequence</subject><subject>Biological and medical sciences</subject><subject>Biosensing Techniques - methods</subject><subject>Biosensors</subject><subject>Biotechnology</subject><subject>Chemistry</subject><subject>Deoxyribonucleic acid</subject><subject>Digoxigenin - chemistry</subject><subject>Digoxigenin - metabolism</subject><subject>DNA</subject><subject>DNA Primers - genetics</subject><subject>DNA Primers - metabolism</subject><subject>DNA, Bacterial - chemistry</subject><subject>DNA, Bacterial - genetics</subject><subject>DNA, Bacterial - metabolism</subject><subject>Electrochemical methods</subject><subject>Electrochemistry</subject><subject>Electrodes</subject><subject>Exact sciences and technology</subject><subject>Food Analysis - methods</subject><subject>Food Microbiology</subject><subject>Food safety</subject><subject>Fundamental and applied biological sciences. 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Technologies</subject><subject>Nanocomposites</subject><subject>Nanocomposites - chemistry</subject><subject>Nucleic Acid Hybridization</subject><subject>Polymerase Chain Reaction</subject><subject>Salmonella</subject><subject>Salmonella - genetics</subject><subject>Salmonella - isolation & purification</subject><subject>Sulfhydryl Compounds - chemistry</subject><subject>Surface Properties</subject><subject>Time Factors</subject><subject>Various methods and equipments</subject><issn>0003-2700</issn><issn>1520-6882</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2009</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpl0MFuEzEQBmALUdFQOPACyELiwGHL2LuxvUcU2oBUtRXNfTVrjxNXm3WwN0J5Dl4YV42aAwdrLp_-Gf-MfRBwKUCKr2gNCF2r_hWbibmEShkjX7MZANSV1ADn7G3OjwBCgFBv2LlohYa5NDP293vc9wNVK1yvw7jm93E4bClhJr7YYBj5L0I7hTjyP2HacOSrTYgDTuT4fQpFchwdvxrITinaDW2DxYEvaYyZxvyU2Jcsx0vAMg6O3-IYbdzuYg4T8YdiYuK-vOsYHX9AT9PhHTvzOGR6f5wXbHV9tVr8qG7ulj8X324qrDVMlTNeCLQe5rqunWraBrQzvdEN9B6k7CW1LdlWIZBqpG8UzaXyHqVysjb1Bfv0HLtL8fee8tQ9xn0ay8ZOCm1Ma2RT0JdnZFPMOZHvduXbmA6dgO6p_O6l_GI_HgP3_ZbcSR7bLuDzEWAuNfmEow35xUkhdCOEOjm0-XTU_wv_AaOamOE</recordid><startdate>20090215</startdate><enddate>20090215</enddate><creator>Brasil de Oliveira Marques, Paulo R</creator><creator>Lermo, Anabel</creator><creator>Campoy, Susana</creator><creator>Yamanaka, Hideko</creator><creator>Barbé, Jordi</creator><creator>Alegret, Salvador</creator><creator>Pividori, M. Isabel</creator><general>American Chemical Society</general><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></search><sort><creationdate>20090215</creationdate><title>Double-Tagging Polymerase Chain Reaction with a Thiolated Primer and Electrochemical Genosensing based on Gold Nanocomposite Sensor for Food Safety</title><author>Brasil de Oliveira Marques, Paulo R ; Lermo, Anabel ; Campoy, Susana ; Yamanaka, Hideko ; Barbé, Jordi ; Alegret, Salvador ; Pividori, M. 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Psychology</topic><topic>Genetic engineering</topic><topic>Genetic technics</topic><topic>Genome, Bacterial</topic><topic>Gold</topic><topic>Gold - chemistry</topic><topic>In vitro gene amplification. Pcr technique</topic><topic>Methods. Procedures. Technologies</topic><topic>Nanocomposites</topic><topic>Nanocomposites - chemistry</topic><topic>Nucleic Acid Hybridization</topic><topic>Polymerase Chain Reaction</topic><topic>Salmonella</topic><topic>Salmonella - genetics</topic><topic>Salmonella - isolation & purification</topic><topic>Sulfhydryl Compounds - chemistry</topic><topic>Surface Properties</topic><topic>Time Factors</topic><topic>Various methods and equipments</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Brasil de Oliveira Marques, Paulo R</creatorcontrib><creatorcontrib>Lermo, Anabel</creatorcontrib><creatorcontrib>Campoy, Susana</creatorcontrib><creatorcontrib>Yamanaka, Hideko</creatorcontrib><creatorcontrib>Barbé, Jordi</creatorcontrib><creatorcontrib>Alegret, Salvador</creatorcontrib><creatorcontrib>Pividori, M. 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Isabel</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Double-Tagging Polymerase Chain Reaction with a Thiolated Primer and Electrochemical Genosensing based on Gold Nanocomposite Sensor for Food Safety</atitle><jtitle>Analytical chemistry (Washington)</jtitle><addtitle>Anal. Chem</addtitle><date>2009-02-15</date><risdate>2009</risdate><volume>81</volume><issue>4</issue><spage>1332</spage><epage>1339</epage><pages>1332-1339</pages><issn>0003-2700</issn><eissn>1520-6882</eissn><coden>ANCHAM</coden><abstract>A novel material for electrochemical biosensing based on rigid conducting gold nanocomposite (nano-AuGEC) is presented. Islands of chemisorbing material (gold nanoparticles) surrounded by nonreactive, rigid, and conducting graphite epoxy composite are thus achieved to avoid the stringent control of surface coverage parameters required during immobilization of thiolated oligos in continuous gold surfaces. The spatial resolution of the immobilized thiolated DNA was easily controlled by merely varying the percentage of gold nanoparticles in the composition of the composite. As low as 9 fmol (60 pM) of synthetic DNA were detected in hybridization experiments when using a thiolated probe. Moreover, for the first time a double tagging PCR strategy was performed with a thiolated primer for the detection of Salmonella sp., one of the most important foodborne pathogens affecting food safety. This assay was performed by double-labeling the amplicon during the PCR with a −DIG and −SH set of labeled primers. The thiolated end allows the immobilization of the amplicon on the nano-AuGEC electrode, while digoxigenin allows the electrochemical detection with the antiDIG-HRP reporter in the femtomole range. Rigid conducting gold nanocomposite represents a good material for the improved and oriented immobilization of biomolecules with excellent transducing properties for the construction of a wide range of electrochemical biosensors such as immunosensors, genosensors, and enzymosensors.</abstract><cop>Washington, DC</cop><pub>American Chemical Society</pub><pmid>19170528</pmid><doi>10.1021/ac801736b</doi><tpages>8</tpages></addata></record> |
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| ispartof | Analytical chemistry (Washington), 2009-02, Vol.81 (4), p.1332-1339 |
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| subjects | Analytical chemistry Animals Base Sequence Biological and medical sciences Biosensing Techniques - methods Biosensors Biotechnology Chemistry Deoxyribonucleic acid Digoxigenin - chemistry Digoxigenin - metabolism DNA DNA Primers - genetics DNA Primers - metabolism DNA, Bacterial - chemistry DNA, Bacterial - genetics DNA, Bacterial - metabolism Electrochemical methods Electrochemistry Electrodes Exact sciences and technology Food Analysis - methods Food Microbiology Food safety Fundamental and applied biological sciences. Psychology Genetic engineering Genetic technics Genome, Bacterial Gold Gold - chemistry In vitro gene amplification. Pcr technique Methods. Procedures. Technologies Nanocomposites Nanocomposites - chemistry Nucleic Acid Hybridization Polymerase Chain Reaction Salmonella Salmonella - genetics Salmonella - isolation & purification Sulfhydryl Compounds - chemistry Surface Properties Time Factors Various methods and equipments |
| title | Double-Tagging Polymerase Chain Reaction with a Thiolated Primer and Electrochemical Genosensing based on Gold Nanocomposite Sensor for Food Safety |
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