Estrogen receptor binding assay of chemicals with a surface plasmon resonance biosensor

We have developed a simple assay method for the evaluation of estrogen receptor (ER) binding capacity of chemicals without the use of radio- or fluorescence-labeled compounds. We used the solution competition assay by the BIACORE biosensor, a surface plasmon resonance biosensor, with estradiol as a ligand, human recombinant ERalpha (hrERalpha) as a high molecular weight (hmw) interactant and test chemicals as analytes. For the ligand, aminated estradiol with a spacer molecule (E2-17PeNH) was synthesized and immobilized on a carboxymethyl dextran-coated sensor chip by the amine coupling method. The injection of the hmw interactant hrERalpha to the biosensor raised the sensorgram, indicating its binding to the ligand E2-17PeNH. The binding of test chemicals to hrERalpha was determined as a reduction in the hrERalpha binding to E2-17PeNH. The dissociation constant for the binding to hrERalpha was calculated for estrone (4.29 x 10(-9) M), estradiol (4.04 x 10(-10) M), estriol (8.35 x 10(-10) M), tamoxifen (2.16 x 10(-8) M), diethylstilbestrol (1.46 x 10(-1)0 M), bisphenot A (1.35 x 10(-6) M) and 4-nonylphenol (7.49 x 10(-6) M), by plotting the data according to an equation based on mass action law. This method can also be used as a high throughput screening method. (C) 2002 Elsevier Science Ltd. All rights reserved.