Classical dot-blot format implemented as an amperometric hybridisation genosensor

Biosens Bioelectron. 2001 Dec;16(9-12):1133-42. doi: 10.1016/s0956-5663(01)00242-1.

Abstract

A new electrochemical hybridisation genosensor has been designed. This genosensor is based on a concept adapted from classical dot-blot DNA analysis, but implemented in an electrochemical biosensor configuration. The use of amperometric transduction and the enzyme label method--that increases the genosensor sensitivity--are the main features of this new approach. The analytical procedure consists of five steps: DNA target immobilisation by adsorption onto a nylon membrane, hybridisation between DNA target and biotin-DNA probe, complexation reaction between biotin-DNA probe and an enzyme (horseradish peroxidase) streptavidin conjugate; integration of the modified membrane onto an electrochemical transducer; and finally, amperometric detection using a suitable substrate for the enzyme labelled duplex. Besides the adapted dot-blot format, a competitive assay in which the target is in solution is reported as well. This procedure, based on amperometric transduction, represents certain advantages with respect to dot-blot analysis: labelled hybrid detection is far simpler, quicker and requires more ordinary or simple reactives; the response obtained is a direct analytical signal via low-cost instrumentation, a nonisotopic labelling is used, and the membranes can be reused. These characteristics are ideal in implementing the procedure developed in kit form.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adsorption
  • Base Sequence
  • Biosensing Techniques / methods*
  • Biotin
  • DNA / analysis*
  • DNA / genetics*
  • DNA, Single-Stranded
  • Electrochemistry
  • Genetic Techniques
  • Horseradish Peroxidase
  • Immunoblotting / methods*
  • Nucleic Acid Hybridization
  • Oligonucleotide Probes
  • Streptavidin

Substances

  • DNA, Single-Stranded
  • Oligonucleotide Probes
  • Biotin
  • DNA
  • Streptavidin
  • Horseradish Peroxidase