Estradiol competition assay

Most of the assays now carried out with isoluminol derivatives are solid-phase competitive assays (Fig. 2), in which the analyte to be determined competes with the labeled analyte for the available binding sites on antibodies that are immobilized on the solid phase. Thus, plastic microspheres have been used to immobilize antibodies for estriol (K12) or thyroxine (W9). Estradiol antibodies have been immobilized onto plastic tubes (K7) or beads (K16). In all of the above cases, the amount of immunoconjugated hormone label is inversely proportional to the amount of free hormone in the analytical sample. Unconjugated labels are removed by decantation or aspiration from the solid phase, and the specific, immunoconjugated labels are measured in a luminometer. This approach has been successfully applied to progesterone analyses in either serum (D6) or saliva (D5). A review of several separation-based assays with isoluminol analogs was presented a few years ago by Kohen et al. (K18). 

Monodisperse microspheres imprinted with theophylline or 17 (3-estradiol were used in competitive radioimmunoassays showing the MIP s high selectivity for the template molecule. In this case the assay is based on the competition of the target molecule with its radioactively labeled analogue for a limited number of antibody binding sites [77,118]. Figure 15 demonstrates that displacing the radioactively marked theophylline from the imprinted polymer was only possible with theophylline as competitor. Structurally related molecules showed effects solely at elevated concentrations [77]. 

The nonlabelled estrogens bound to receptor in the test sample will be competitively displaced by the addition of 17-P-estradiol labelled with tritium for radiochemical assay. 

In our previous studies, DBM showed inhibitory effect on DMBA-induced covalent binding of [ H]DMBA to mammary gland DNA. Therefore, we subsequently evaluated its influence on hormonal mitogenic action toward tumor promotion. Because of the structural similarity between estradiol (Ea) and DBM, a competitive estrogen-receptor binding assay was used in our studies to examine the inhibitory effects of DBM on estradiol-dependent action. 

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