ELECTROCHEMICAL IMMUNOASSAY - SIMPLE KINETIC DETECTION OF ALKALINE-PHOSPHATASE ENZYME LABELS IN LIMITED AND EXCESS REAGENT SYSTEMS

Kinetic measurement of an alkaline phospharase enzyme label was perfor med with amperometric detection using conversion of substrate 4-aminop henyl phosphate to the electroactive phenolic, measured at +125 to +20 0 mV (vs. Ag/AgCl). Calibration plots for thyroxine-binding globulin ( TBG) and cortisol were obtained by excess reagent and limited reagent immunoassays, respectively, which were comparable with those obtained using colorimetric detection. These provided a measurement range of 31 -1000 mu g/L for TBG and 100-2000 nM for cortisol. An antibody-bound l abel was monitored in this system, and covering the electrode with a s imple dialysis or polycarbonate membrane avoided electrode fouling. Th e kinetic mode of detection reduced assay time and helped to simplify the assay protocol.