title: Rapid amperometric verification of PCR amplification of DNA.
abstract: Amplification of an 800-base template was verified in a 10-min test on a 2-microL sample of the PCR product solution. For verification, digoxigeninylated primers and biotinylated d-UTP-16-biotin were added to the amplification solution. The resulting amplified product was digoxigeninlabeled at its 3'-end and was also labeled with multiple biotin functions along its chain. The detecting electrode was coated with an electron-conducting redox hydrogel to which anti-digoxin monoclonal antibody was covalently bound. The amplified DNA was captured by the electrode through conjugation of its 3'-digoxigenin with the antibody. Exposure to a solution of horseradish peroxidase-labeled avidin led to capture of the enzyme and switched the redox hydrogel from a noncatalyst to catalyst for H2O2 electroreduction. The switching resulted in an H2O2 electroreduction current density of 2.1 +/- 0.9 microA cm-2 in 10-4 M H2O2 at Ag/AgCl potential and at 25 degrees C.
authors.affiliation: Department of Chemical Engineering, University of Texas at Austin 78712-1062, USA.
authors.forename: T
authors.initials: T
authors.lastname: de Lumley-Woodyear
authors.forename: C N
authors.initials: CN
authors.lastname: Campbell
authors.forename: E
authors.initials: E
authors.lastname: Freeman
authors.forename: A
authors.initials: A
authors.lastname: Freeman
authors.forename: G
authors.initials: G
authors.lastname: Georgiou
authors.forename: A
authors.initials: A
authors.lastname: Heller
mesh_terms: D004247:DNA / Q000032:analysis*; D004563:Electrochemistry / Q000379:methods; D005784:Gene Amplification; D016133:Polymerase Chain Reaction / Q000379:methods*