Brasil
| Otro
Polysiloxane-poly(propylene oxide) hybrid discs as solid phase in anti-HCV detection using a recombinant core protein
| dc.contributor | Universidade Estadual Paulista (UNESP) | |
| dc.creator | Taghavini, S. A. | |
| dc.creator | Mikawa, A. Y. | |
| dc.creator | Yamanaka, H. | |
| dc.creator | Henrique-Silva, F. | |
| dc.creator | Costa, P. I. | |
| dc.date | 2014-05-20T15:31:39Z | |
| dc.date | 2016-10-25T18:07:33Z | |
| dc.date | 2014-05-20T15:31:39Z | |
| dc.date | 2016-10-25T18:07:33Z | |
| dc.date | 2008-04-15 | |
| dc.date.accessioned | 2017-04-06T00:23:17Z | |
| dc.date.available | 2017-04-06T00:23:17Z | |
| dc.identifier | Talanta. Amsterdam: Elsevier B.V., v. 75, n. 2, p. 461-465, 2008. | |
| dc.identifier | 0039-9140 | |
| dc.identifier | http://hdl.handle.net/11449/40724 | |
| dc.identifier | http://acervodigital.unesp.br/handle/11449/40724 | |
| dc.identifier | 10.1016/j.talanta.2007.11.036 | |
| dc.identifier | WOS:000255270700021 | |
| dc.identifier | http://dx.doi.org/10.1016/j.talanta.2007.11.036 | |
| dc.identifier.uri | http://repositorioslatinoamericanos.uchile.cl/handle/2250/883473 | |
| dc.description | In 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. | |
| dc.language | eng | |
| dc.publisher | Elsevier B.V. | |
| dc.relation | Talanta | |
| dc.rights | info:eu-repo/semantics/closedAccess | |
| dc.subject | hepatitis C virus | |
| dc.subject | core protein | |
| dc.subject | recombinant antigen | |
| dc.subject | siloxane-poly(propylene oxide) discs | |
| dc.title | Polysiloxane-poly(propylene oxide) hybrid discs as solid phase in anti-HCV detection using a recombinant core protein | |
| dc.type | Otro |