Simple enzyme-amplified amperometric detection of a 38-base oligonucleotide at 20 pmol L(-1) concentration in a 30- microL droplet

A 38-base DNA sequence has been detected at 20 pmol L(-1) concentration in 15-35- microL droplets by means of an electrochemical enzyme-amplified sandwich-type assay on a mass-manufacturable screen-printed carbon electrode. Formation of the sandwich brought the horseradish peroxidase-label of the detection sequence into electrical contact with a pre-electrodeposited redox polymer, making the sandwich an electrocatalyst for the reduction of hydrogen peroxide to water at +0.2 V (Ag/AgCl). Sensitivity twenty times better than that of a related system resulted from: 1. fivefold reduction of the noise by substituting the formerly used poly( N-vinyl imidazole)-co-acrylamide comprising redox co-polymer with poly(4-vinyl pyridine)-co-acrylamide comprising redox polymer, enabling use of the electrodes at a more oxidizing potential at which noise (the rate of non-enzyme catalyzed electroreduction currents of dissolved oxygen and hydrogen peroxide) was lower; 2. doubling of the catalytic electroreduction current upon electrodeposition of a second layer of the redox polymer on the capture sequence-containing film; and 3. doubling of the current by increasing the coverage by the capture sequence.