Rapid amperometric verification of PCR amplification of DNA

Amplification of an 800-base template was verified in a 10-min test on a 2- mu L sample of the PCR product solution. For verification, digoxigeninylate d 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, Th ee detecting electrode was coated with an electron-conducting redox hydroge l to which anti-digoxin monoclonal antibody was covalently bound. The ampli fied DNA was captured by the electrode through conjugation of its 3'-digoxi genin with the antibody. Exposure to a solution of horseradish peroxidase-l abeled avidin led to capture of the enzyme and switched the redox hydrogel from a noncatalyst to a catalyst for H2O2 electroreduction. The switching r esulted in an H2O2 electroreduction current density of 2.1 +/- 0.9 mu A cm( -2) in 10(-4) M H2O2 at Ag/AgCl potential and at 25 degrees C.