Elimination of 'hook-effect' in two-site immunoradiometric assays by kinetic rate analysis

Abstract

Two-site or sandwich immunoradiometric assays (IRMAs) offer theoretical advantages over competitive immunoassay systems for sensitivity, precision, and rapid incubation. The practical realization of these advantages has been limited by the phenomenon of the 'high-dose hook effect,' such that high concentrations of an analyte give similar responses to those of much lower concentrations. We have developed a kinetic rate monitoring IRMA system for use with the Kineti-Count 48(TM), an automated kinetic radioassay analyzer, which eliminates 'hook-effect' interference, thereby permitting optimal assay design for increasing sensitivity and reducing incubation time. Practical illustration of these concepts is demonstrated by a 10-min, automated, quantitative assay we developed for human choriogonadotropin. The assay can detect as little as 1.2 int. units/L and kinetically screens for the hook effect. Kinetic rate analysis of the two-site IRMA and potentially of nonisotopic counterparts permits improvements in the speed and reliability of these immunoassays.