Homogeneous two-site immunometric assay kinetics as a theoretical tool for data analysis.

The easily accessible kinetics of a new homogeneous two-site fluorometric immunoassay for prolactin was studied, in order to determine its usefulness for assay data reduction and optimization. The combined use of a simple descriptive model fitted to experimental data and a mechanistic model to simulate the kinetics revealed that (i) the kinetics curve presented an early inflexion point. Its time of occurrence was constant as long as the antigen concentration was below the smallest antibody concentration and decreased to zero for higher concentrations. It may therefore be used as an indicator of hooked samples. (ii) The kinetics steepest slope was correlated with antigen concentration. Its use as a dose-response curve variable would allow higher concentrations to be assayed than with the classical end-point dose-response curve. The results suggest that control and exploitation of kinetic parameters could help to improve the rapidity, analytical range, and reliability of homogeneous two-site immunometric assays.