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-microm gaps and 2.4-microm 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-microL drop was detectable and the immunoassay was complete within 10 min with a detection limit of 3.5x10(-15)mol mouse IgG.
Thomas
Jennifer H
JH
Department of Chemistry, University of Cincinnati, P.O. Box 210172, Cincinnati, OH 45221-0172, USA.
Kim
Sang Kyung
SK
Hesketh
Peter J
PJ
Halsall
H Brian
HB
Heineman
William R
WR
D000627:Aminophenols / Q000032:analysis; D000818:Animals; D002138:Calibration; D004563:Electrochemistry / Q000379:methods*; D004566:Electrodes; D007097:Imines / Q000737:chemistry; D007118:Immunoassay / Q000379:methods*; D007124:Immunoenzyme Techniques; D007074:Immunoglobulin G / Q000032:analysis*; D051379:Mice; D008832:Microchemistry; D010084:Oxidation-Reduction; D011809:Quinones / Q000737:chemistry; D013997:Time Factors; D001616:beta-Galactosidase / Q000737:chemistry / Q000378:metabolism
D000627:Aminophenols; D007097:Imines; D007074:Immunoglobulin G; D011809:Quinones; D001616:beta-Galactosidase