This study deals with the development of enantioselective flow-through immunosensors for triiodothyronine (T3) and tetraiodothyronine (thyroxine, T4) on the basis of a competitive assay using enantioselective antibodies. The instrumental set-up is based o

This study deals with the development of enantioselective flow-through immunosensors for triiodothyronine (T3) and tetraiodothyronine (thyroxine, T4) on the basis of a competitive assay using enantioselective antibodies. The instrumental set-up is based on a simple sequential-injection system equipped with a chemiluminescence detector and an immunoreactor, which consists of a flow-cell packed with immobilized haptens. As haptens, 4-amino-l-phenylalanine (4-amino-l-Phe), 4-amino-d-Phe or l-T3 were used. Antibodies directed against 4-amino-l- or d-Phe or l-T3 were labeled with an acridinium ester. Three different approaches for immobilizing the haptens were investigated including simple adsorption on polystyrene, chemical binding to an activated methacrylate polymer and binding via the biotin–streptavidin binding (BSB) system. The latter approach showed the best results regarding repeatability and sensivity. Using biotinylated l-T3 immobilized onto a streptavidin-derivatized trisacryl support and labeled anti-l-T3 antibodies, a detection limit of 15.5 fmol/ml for l-T3 was obtained. One assay cycle including regeneration takes only about 5 min. This approach was applied to detect l-T3 in plasma samples without any sample pre-treatment. The average recovery from spiked plasma sample was about 93% with a R.S.D. below 5%.