Microbead-based electrochemical immunoassay with interdigitated array electrodes

The objective of this study was to develop a sensitive and miniaturized immunoassay by coupling a microbead-based immunoassay with an interdigitated array (IDA) electrode. An IDA electrode amplifies the signal by recycling an electrochemically redox-reversible molecule. The microfabricated platinum electrodes had 25 pairs of electrodes with 1.6-mum gaps and 2.4-mum widths. An enzyme-labeled sandwich immunoassay on paramagnetic microbeads with mouse IgG as the analyte and beta-galactosidase as the enzyme label was used as the model system. beta-Galactosidase converted p-aminophenyl beta-D-galactopyranoside to p-aminophenol (PAP). This enzyme reaction was measured continuously by positioning the microbeads near the electrode surface with a magnet. Electrochemical recycling occurred with PAP oxidation to p-quinone imine (PQI) at +290 mV followed by PQI reduction to PAP at -300 mV vs Ag/AgCl. Dual-electrode detection amplified the signal fourfold compared to single-electrode detection, and the recycling efficiency reached 87%. A calibration curve of PAP concentration vs anodic current was linear between 10(-4) and 10(-6) M. A signal from 1000 beads in a 20-muL drop was detectable and the immunoassay was complete within 10 min with a detection limit of 3.5 x 10(-15) mol mouse IgG. (C) 2004 Elsevier Inc. All rights reserved.