Abstract
Fluorescein, antifluorescein reactions have been studied by initial fluorescence polarization and intensity rate methods. Novel rapid stopped flow instrumentation (msec time range) as well as slower but more sensitive instrumentation has yielded second order rate constants over wide ranges in concentration and time. A bimolecular mechanism for the fluorescein, univalent antifluorescein system has been proposed in which two types of site interactions occur. This mechanism accounts for the fact that fluorescence polarization data consistently yield second order rate constants which are a factor of 10 higher than those obtained from fluorescence quenching data. Comparisons of kinetic behavior of free fluorescein and of fluorescein covalently attached to a normal IgG carrier reacting with Fab antifluorescein have suggested a “carrier effect” of a steric nature. The large equilibrium association constant for fluorescein, antifluorescein systems has been found to be due to extremely small values of the back reaction rate constants; i. e. 10 −3 to 10 −4 as great as those reported for DNP, anti DNP systems.
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