Immunoassays and kits for use therein which include low affinity

Abstract

An immunoassay for assaying an antigenic substance (Ag) in a fluid. The immunoassay is the type which comprises contacting the fluid with at least one first entity selected from a group consisting of an antibody (Ab) to the Ag, a soluble, labeled antibody (L-Ab.sub.a) to the Ag, and an antibody (Ab.sub.b) to the Ag bound to a solid support (SC). The immunoassay is characterized in that the fluid is contacted with at least one additional entity selected from a group consisting of at least one different type of soluble, labeled antibody (L-Ab.sub.c) to the Ag, at least one different type of antibody (Ab.sub.d) bound to a solid carrier (SC.sub.1), and at least one different type of antibody (Ab.sub.e) to the Ag. The SC.sub.1 is selected from a group consisting of SC, at least one different solid carrier (SC.sub.2), and mixtures thereof (SC and SC.sub.2). Each type of L-Ab.sub.c, Ab.sub.d -SC.sub.1, and Ab.sub.e has a lower average affinity constant (K) for Ag than each respective K of L-Ab.sub.a, Ab.sub.b -SC and Ab.sub. e ; and the additional entity is present in an amount sufficient to avoid a hook effect. Also, a reagent of the type comprising at least one first entity selected from a group consisting of L-Ab.sub.a, Ab.sub.b -SC, and Ab. The reagent is characterized in that it further comprises at least one additional entity selected from a group consisting of at least one different type of L-Ab.sub.c and at least one different type of Ab.sub.d -SC.sub.1, and at least one different type of Ab.sub.e. Ab, L-Ab.sub.a, Ab.sub.b -SC, L-Ab.sub.c, Ab.sub.d -SC.sub.1, and Ab.sub.e are as defined above.