Antibodies conjugated with new highly luminescent Eu3+ and Tb3+ chelates as markers for time resolved immunoassays. Application to simultaneous determination of clenbuterol and free cortisol in horse urine

Highly luminescent Eu3+ and Tb3+ complexes of 10-[4-(3-isothiocyanatopropoxy)benzoylmethyl]-1,4,7,10-tetraazacyclodode cane-1,4,7 triacetic acid Eu3+ subset of 1 and Tb3+ subset of 1 were conjugated with a goat anti-rabbit IgG and a rabbit anti-mouse IgG, respectively, and applied as markers in a time resolved immunoassay for simultaneous quantitative determination of anabolic compounds clenbuterol (CL) and hydrocortisone (HC). The assay was performed in horse urine, using a monoclonal antibody specific to CL and a rabbit polyclonal antibody specific to the free HC. These lanthanide chelates are very stable and highly luminescent in aqueous solution and allowed to reach 10 mu g L-1 and 40 mu g L-1 sensitivities for CL and for HC, respectively. Application to the horse urine, that is a very complex matrix, has a considerable interest in the control of illegal use of these compounds.