Estrogen receptor in very small breast tumor specimens: a modified charcoal-gelatin assay

We described recently a modified charcoal-gelatin (MCG) assay for measuring progesterone receptor activity in low-protein cytosols. We showed that pre-mixing of gelatin with sample cytosols (final gelatin concentration 0.1%) and removal of unbound steroid by a 1% charcoal suspension with 0.1% gelatin but without dextran preserves the progesterone receptor activity in dilute cytosol. For estrogen receptor (ER), as for progesterone receptor, the efficiency of the standard dextran-coated charcoal (DCC) assay drops rapidly as samples are diluted much below 1 mg protein per ml. We have therefore applied the MCG procedure to the assay of ER in breast tumor cytosols. We find that MCG is far more efficient for ER at low protein concentrations than either the DCC method or three other methods recommended previously for dilute samples, retaining at least 60% efficiency even at 0.01 mg protein per ml. The measured Kd of the receptor for estradiol is the same by MCG as by DCC. A series of human breast tumor biopsies assayed by MCG at 0.1 mg protein per ml gave about the same ER values (fmol/mg) as at 1 mg/ml, while the DCC efficiency for ER at the lower concentration averaged only 32%. In combination with 125I-labeled estradiol, this MCG method should allow accurate ER assays of extremely small breast cancer specimens.