Genosensor on gold films with enzymatic electrochemical detection of a SARS virus sequence.
Identifieur interne : 002815 ( PubMed/Corpus ); précédent : 002814; suivant : 002816Genosensor on gold films with enzymatic electrochemical detection of a SARS virus sequence.
Auteurs : Patricia Abad-Valle ; M Teresa Fernández-Abedul ; Agustín Costa-GarcíaSource :
- Biosensors & bioelectronics [ 0956-5663 ] ; 2005.
English descriptors
- KwdEn :
- Biosensing Techniques (instrumentation), Biosensing Techniques (methods), DNA, Viral (analysis), DNA, Viral (genetics), Electrochemistry (instrumentation), Electrochemistry (methods), Equipment Design, Equipment Failure Analysis, Gold (chemistry), In Situ Hybridization (instrumentation), In Situ Hybridization (methods), SARS Virus (genetics), SARS Virus (isolation & purification), Sequence Analysis, DNA (instrumentation), Sequence Analysis, DNA (methods).
- MESH :
- chemical , analysis : DNA, Viral.
- chemical , chemistry : Gold.
- chemical , genetics : DNA, Viral.
- genetics : SARS Virus.
- instrumentation : Biosensing Techniques, Electrochemistry, In Situ Hybridization, Sequence Analysis, DNA.
- isolation & purification : SARS Virus.
- methods : Biosensing Techniques, Electrochemistry, In Situ Hybridization, Sequence Analysis, DNA.
- Equipment Design, Equipment Failure Analysis.
Abstract
A hybridisation-based genosensor was designed on a 100 nm sputtered gold film. This material worked as an immobilisation and transduction surface. A 30-mer sequence that encodes a short lysine-rich region, unique to SARS (severe acute respiratory syndrome) virus, was chosen as target. A complementary strand (probe), labelled with a thiol group at the 3'-end, was immobilised on the film. After blocking the surface, hybridisation with the biotin-conjugated SARS strand (at the 3'-end) took place. Interaction with alkaline phosphatase-labelled streptavidin permits amplified indirect electrochemical detection. The analytical signal is constituted by an electrochemical process of indigo carmine, the soluble product of the enzymatic hydrolysis of 3-indoxyl phosphate. The use of a sensitive electrochemical technique such as square wave voltammetry allowed a detection limit of 6 pM to be obtained for this DNA sequence, lower than any other found in the bibliography. The parameters affecting the methodology were studied, with special attention being placed on selectivity. Specificity was clearly enhanced when interaction time and stringency (in the form of formamide percentage) were increased. With 1h of strand interaction and employing 50% of formamide in the hybridisation buffer, a 3-base mismatch strand was perfectly distinguished from the complementary.
DOI: 10.1016/j.bios.2004.10.019
PubMed: 15797323
Links to Exploration step
pubmed:15797323Le document en format XML
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This material worked as an immobilisation and transduction surface. A 30-mer sequence that encodes a short lysine-rich region, unique to SARS (severe acute respiratory syndrome) virus, was chosen as target. A complementary strand (probe), labelled with a thiol group at the 3'-end, was immobilised on the film. After blocking the surface, hybridisation with the biotin-conjugated SARS strand (at the 3'-end) took place. Interaction with alkaline phosphatase-labelled streptavidin permits amplified indirect electrochemical detection. The analytical signal is constituted by an electrochemical process of indigo carmine, the soluble product of the enzymatic hydrolysis of 3-indoxyl phosphate. The use of a sensitive electrochemical technique such as square wave voltammetry allowed a detection limit of 6 pM to be obtained for this DNA sequence, lower than any other found in the bibliography. The parameters affecting the methodology were studied, with special attention being placed on selectivity. Specificity was clearly enhanced when interaction time and stringency (in the form of formamide percentage) were increased. With 1h of strand interaction and employing 50% of formamide in the hybridisation buffer, a 3-base mismatch strand was perfectly distinguished from the complementary.</div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">15797323</PMID><DateCompleted><Year>2005</Year><Month>08</Month><Day>01</Day></DateCompleted><DateRevised><Year>2020</Year><Month>04</Month><Day>09</Day></DateRevised><Article PubModel="Print"><Journal><ISSN IssnType="Print">0956-5663</ISSN><JournalIssue CitedMedium="Print"><Volume>20</Volume><Issue>11</Issue><PubDate><Year>2005</Year><Month>May</Month><Day>15</Day></PubDate></JournalIssue><Title>Biosensors & bioelectronics</Title><ISOAbbreviation>Biosens Bioelectron</ISOAbbreviation></Journal><ArticleTitle>Genosensor on gold films with enzymatic electrochemical detection of a SARS virus sequence.</ArticleTitle><Pagination><MedlinePgn>2251-60</MedlinePgn></Pagination><Abstract><AbstractText>A hybridisation-based genosensor was designed on a 100 nm sputtered gold film. This material worked as an immobilisation and transduction surface. A 30-mer sequence that encodes a short lysine-rich region, unique to SARS (severe acute respiratory syndrome) virus, was chosen as target. A complementary strand (probe), labelled with a thiol group at the 3'-end, was immobilised on the film. After blocking the surface, hybridisation with the biotin-conjugated SARS strand (at the 3'-end) took place. Interaction with alkaline phosphatase-labelled streptavidin permits amplified indirect electrochemical detection. The analytical signal is constituted by an electrochemical process of indigo carmine, the soluble product of the enzymatic hydrolysis of 3-indoxyl phosphate. The use of a sensitive electrochemical technique such as square wave voltammetry allowed a detection limit of 6 pM to be obtained for this DNA sequence, lower than any other found in the bibliography. The parameters affecting the methodology were studied, with special attention being placed on selectivity. Specificity was clearly enhanced when interaction time and stringency (in the form of formamide percentage) were increased. With 1h of strand interaction and employing 50% of formamide in the hybridisation buffer, a 3-base mismatch strand was perfectly distinguished from the complementary.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Abad-Valle</LastName><ForeName>Patricia</ForeName><Initials>P</Initials><AffiliationInfo><Affiliation>Departamento de Química Física y Analítica, Universidad de Oviedo, Asturias, 33006 Oviedo, Spain.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Fernández-Abedul</LastName><ForeName>M Teresa</ForeName><Initials>MT</Initials></Author><Author ValidYN="Y"><LastName>Costa-García</LastName><ForeName>Agustín</ForeName><Initials>A</Initials></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D023362">Evaluation Study</PublicationType><PublicationType UI="D016428">Journal Article</PublicationType><PublicationType UI="D013485">Research Support, Non-U.S. 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MajorTopicYN="Y">genetics</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D004563" MajorTopicYN="N">Electrochemistry</DescriptorName><QualifierName UI="Q000295" MajorTopicYN="Y">instrumentation</QualifierName><QualifierName UI="Q000379" MajorTopicYN="N">methods</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D004867" MajorTopicYN="N">Equipment Design</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D019544" MajorTopicYN="N">Equipment Failure Analysis</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D006046" MajorTopicYN="N">Gold</DescriptorName><QualifierName UI="Q000737" MajorTopicYN="N">chemistry</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D017403" MajorTopicYN="N">In Situ Hybridization</DescriptorName><QualifierName UI="Q000295" MajorTopicYN="Y">instrumentation</QualifierName><QualifierName UI="Q000379" MajorTopicYN="N">methods</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D045473" 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