A sensor capable of detecting a specific DNA sequence was designed by bulk modification of a graphite epoxy composite electrode with streptavidin (2% w/w). Streptavidin is used to immobilise a biotinylated capture DNA probe to the surface of the electrode. Simultaneous hybridisation occurs between the biotin DNA capture probe and the target-DNA and between the target-DNA and a digoxigenin modified probe. The rapid binding kinetic of streptavidin-biotin allows a one step immobilisation/hybridisation procedure. Secondly, enzyme labelling of the DNA duplex occurs via an antigen-antibody reaction between the Dig-dsDNA and an anti-Dig-HRP. Finally, electrochemical detection is achieved through a suitable substrate (H2O2) for the enzyme-labelled duplex. Optimisation of the sensor design, the modifier content and the immobilisation and hybridisation times was attained using a simple nucleotide sequence. Regeneration of the surface is achieved with a simple polishing procedure that shows good reproducibility. The generic use of a modified streptavidin carbon-polymer biocomposite electrode capable of surface regeneration and a one step hybridisation/immobilisation procedure are the main advantages of this approach. In DNA analysis, this procedure, if combined with the polymerase chain reaction, would represent certain advantages with respect to classical techniques, which prove to be time consuming in situations where a simple and rapid detection is required. This innovative developed material may be used for the detection of any analyte that can be coupled to the biotin-streptavidin reaction, as is the case of immunoassays. © 2003 Elsevier B.V. All rights reserved.
|Journal||Biosensors and Bioelectronics|
|Publication status||Published - 30 Nov 2003|
- Amperometric genosensor
- Anti-digoxigenin antibody
- DNA hybridisation
- Horseradish peroxidase labelling
- Streptavidin carbon-polymer biocomposite