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dc.contributor.authorTaghavini, S. A. [UNESP]
dc.contributor.authorMikawa, A. Y. [UNESP]
dc.contributor.authorYamanaka, H. [UNESP]
dc.contributor.authorHenrique-Silva, F.
dc.contributor.authorCosta, P. I.
dc.date.accessioned2014-05-20T15:31:39Z
dc.date.available2014-05-20T15:31:39Z
dc.date.issued2008-04-15
dc.identifierhttp://dx.doi.org/10.1016/j.talanta.2007.11.036
dc.identifier.citationTalanta. Amsterdam: Elsevier B.V., v. 75, n. 2, p. 461-465, 2008.
dc.identifier.issn0039-9140
dc.identifier.urihttp://hdl.handle.net/11449/40724
dc.description.abstractIn this work, siloxane-poly(propylene oxide) discs (PPO disc) prepared using the sol-gel process were used as solid phase in enzyme-linked immunosorbent assays (ELISA) for the detection of anti-hepatitis C virus (HCV) antibodies. The HCV RNA from serum (genotype 1b) was submitted to the RT-PCR technique and subsequent amplification of the HCV core 408 pb. This fragment was cloned into expression vector pET42a and expressed in Escherichia coli as recombinant protein with glutathione S-transferase (GST). Cell cultures were grown and induced having a final concentration of 0.4 x 10(-3) mol L-1 of IPTG. After induction, the cells were harvested and the soluble fraction was analyzed using polyacrilamide gel 15% showing a band with an approximate molecular weight of 44 kDa, the expected size for this GST-fused recombinant protein. The recombinant protein was purified and continued by immunological detection using HCV-positive serum and showed no cross-reactivity with positive samples for other infectious diseases. An ELISA was established using 1.25 ng of recombinant protein per PPO disc, a dilution of 1: 10,000 and 1:40 for a peroxidase conjugate and serum, respectively, and solutions of hydrogen peroxide and 3,3',5,5'-tetra-methylbenzidine in a ratio of 1: 1. The proposed methodology was compared with the ELISA conventional polystyrene-plate procedure and the performance of the PPO discs as a matrix for immunodetection gave an easy synthesis, good performance and reproducibility for commercial application. (c) 2007 Elsevier B.V. All rights reserved.en
dc.format.extent461-465
dc.language.isoeng
dc.publisherElsevier B.V.
dc.relation.ispartofTalanta
dc.sourceWeb of Science
dc.subjecthepatitis C virusen
dc.subjectcore proteinen
dc.subjectrecombinant antigenen
dc.subjectsiloxane-poly(propylene oxide) discsen
dc.titlePolysiloxane-poly(propylene oxide) hybrid discs as solid phase in anti-HCV detection using a recombinant core proteinen
dc.typeArtigo
dcterms.licensehttp://www.elsevier.com/about/open-access/open-access-policies/article-posting-policy
dcterms.rightsHolderElsevier B.V.
dc.contributor.institutionUniversidade Estadual Paulista (Unesp)
dc.contributor.institutionUniversidade Federal de São Carlos (UFSCar)
dc.description.affiliationUniv Estadual Paulista, Univ Estadual Paulista, Inst Quim, BR-14800900 São Paulo, Brazil
dc.description.affiliationUNESP, Univ Estadual Paulista, Fac Ciencias Farmaceut, BR-14801902 Araraquara, Brazil
dc.description.affiliationUniversidade Federal de São Carlos (UFSCar), UFSCar, BR-13565905 São Carlos, SP, Brazil
dc.description.affiliationUnespUniv Estadual Paulista, Univ Estadual Paulista, Inst Quim, BR-14800900 São Paulo, Brazil
dc.description.affiliationUnespUNESP, Univ Estadual Paulista, Fac Ciencias Farmaceut, BR-14801902 Araraquara, Brazil
dc.identifier.doi10.1016/j.talanta.2007.11.036
dc.identifier.wosWOS:000255270700021
dc.rights.accessRightsAcesso restrito
dc.identifier.lattes6720223715917381
dc.identifier.orcid0000-0002-3350-8308
unesp.author.lattes6720223715917381[5]
unesp.author.orcid0000-0002-3350-8308[5]
dc.relation.ispartofjcr4.244
dc.relation.ispartofsjr1,186
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