Rotating disk electrode amperometric detection for a bead-based immunoassay

Abstract

Amperometric detection with a rotating disk electrode (RDE) for a bead based enzyme immunoassay for the model antigen mouse IgG has been developed. Alkaline phosphatase was used as the enzyme label for the conversion of the substrate 4-aminophenyl phosphate to the electroactive 4-aminophenol (PAP). The detection scheme allows enzyme turnover to be monitored continuously and thus reduces errors associated with obtaining data at fixed times as in flow injection analysis. The determination of PAP by the RDE has a linear range from 5×10−9 to 5×10−5 M. An 800 μl electrochemical cell was used to minimize enzyme product dilution. A linear response for mouse IgG was obtained in this non-optimized system over the concentration range of 50–5000 ng ml−1.