Endocrinology, Vol 106, 56-60, Copyright © 1980 by Endocrine Society

ARTICLES

A simple modification of the estrogen receptor exchange assay: validation in nuclei from the rat uterus and a mouse mammary tumor

DR Schoenberg and JH Clark

Crude nuclear preparations from the rat uterus and the MXT mouse mammary tumor contain endogenous nucleases whose activity results in the loss of estrogen-binding sites from these fractions due to DNA breakdown. Although this is not a significant problem for studies in the uterus, the majority of the receptors present in nuclei from the tumor were lost from the particulate fraction under the standard conditions for the [3H]estradiol exchange assay. The addition of 5 mM Ca++ to either of these nuclear preparations resulted in the aggregation of the released estrogen receptors with particulate components and, hence, a total inhibition of receptor release into the soluble (wash) fractions. Ca++ had no effect on the affinity of the receptors for [3H]estradiol, although in some cases it slowed the overall rate of binding. The addition of Ca++ to the nuclear estrogen exchange assay may therefore prove to be a simple yet powerful aide in studies on the potential hormonal responsiveness of human and experimental mammary tumors.