In vitro analysis of xenoestrogens by enzyme linked receptor assays (ELRA)

Receptor binding assays are a modern approach to the effects-related analysis of xenoendocrines. Human estrogen receptors are used to develop an analytical tool for the detection of estrogenic substances in environmental samples. A sensitive Enzyme Linked Receptor Assay (ELRA) was developed in a microwell plate format. The receptor assay is based on similar principles as competitive immunoassays (ELISA). However, receptor binding always implies a biological effect, either agonistic or antagonistic. The choice of suitable tracers or hapten conjugates is an important step in assay development. For this purpose and for the determination of receptor-affinities of relevant xenoestrogens, a surface plasmon resonance (SPR) biosensor (BIAcore) was used for binding studies with immobilized receptors. Results with commercially available hapten-conjugates (estradiol-BSA) show a direct correlation between the amount of immobilized estradiol receptor and the amount of bound hapten-conjugate. Based on these BIAcore experiments an ELRA was developed. The calibration curves show a detection limit of 0.1 microgram/l for 17 beta-estradiol. Cross-reactivities of different steroids and xenoestrogens are reported.