The inhibition of the proliferative stimulation exercised by estrogens on neoplastic cells is the goal of all endocrine therapies in breast cancer. Under various circumstances, e.g. with the use of aromatase inhibitors, this result can be obtained by blocking the synthetic pathway and, consequently, by lowering the circulating levels of estradiol (E2), estrone (E1), and estrone sulfate (E1-S). The evaluation of these hormones in plasma could therefore represent a useful indicator of the biological efficacy of the therapy. However, the measurement of circulating steroids in a large series of patients is often a complicated procedure. Indirect methods of extraction are time consuming and expensive while the analytical sensitivity of direct methods is not sufficient to measure the residual levels of E2, E1, and E1-S. In this paper we describe a novel extraction method for the evaluation of plasma levels of E2, E1, and E1-S. This new method consists of solid phase extraction followed by a highly specific radioimmunoassay. The sensitivity of the assay is 0.6 pg/ml, 2.0 pg/ml and 7.0 pg/ml for E2, E1, and E1-S, respectively.