Estrogen binding sites in the nucleus of normal and malignant human tissue: optimization of an exchange assay for the measurement of specific binding.

An exchange assay has been validated and used to measure the total concentration of estrogen receptors in the nuclei of cells from human breast tumors and myometrium. Tissue homogenates were centrifuged through 1.2 M sucrose pads to separate crude chromatin from soluble ane membranous fractions. The yield of DNA by this procedure was approximately 80%. Total binding sites were measured by incubating the sedimented pellets with tritiated estradiol at 30 degrees for 60 min and then precipitating the receptor-steroid complexes with protamine sulfate. Saturation analysis by this procedure provides evidence for the presence of a specific, nuclear estrogen-binding site in addition to the established estrogen receptor in both these tissues.