New Approach for Visualizing Estrogen Receptors in Target Cells Using Inherently Fluorescent Ligands and Image Intensification

Pierre M. Martin, Henri P. Magdelenat, Bahia Benyahia, Odile Rigaud, John A. Katzenellenbogen, Pierre M. Martin, Henri P. Magdelenat, Bahia Benyahia, Odile Rigaud, John A. Katzenellenbogen, Pierre M. Martin, Henri P. Magdelenat, Bahia Benyahia, Odile Rigaud, John A. Katzenellenbogen

Research output: Contribution to journalArticlepeer-review

Abstract

Four fluorescent estrogen ligands were investigated as agents for visualization of estrogen receptors in cells: 2-(2,4-dihydroxy-phenyl)-6-hydroxy-3-benzofurancarboxylic add δ-lactone (coumestrol) and 9(11)-dehydro-12-oxoestradiol [12-oxo-1,3,5-(10),9(11)-estratetraene-3,17β-diol] (12-oxoestradiol), which are inherently fluorescent compounds; and tamoxifen {(Z)-1-[4-(2-dimethylaminoethoxy)phenyl]-1,2-diphenyM -butene} and 4-hydroxytamoxifen {(Z)-1 -[4-(2-dimethylaminoethoxy)phenyl]-1 -(4-hydroxyphenyl)-2-phenyl-1 -butene}, which become maximally fluorescent only after ultraviolet irradiation. By conventional fluorescence techniques, these agents can be detected down to 10-8 m in water, but only to 10-6 to 10-7 m in protein solutions; however, by photon-counting spectrofluorimetry, coumestrol and 12-oxoestradiol can be detected in protein solutions down to 5 x 10-10 m. Three of these compounds have good affinity for the estrogen receptor coumestrol (20%); 12-oxoestradiol (12%); and 4-hydroxytamoxifen (37%), relative to estradiol (100%). Under conditions where autoradiographic controls indicate that most of the estrogen receptor of MCF-7 human breast cancer cells is in the nucleus, we could demonstrate nuclear fluorescence using 10-9 m concentrations of coumestrol, 12-oxoestradiol, and 4-hydroxytamoxifen. This nuclear fluorescence was abolished by a 200-fold excess of diethylstilbestrol and could only be observed through a fluorescence microscope equipped with a microchannel image intensifier and a video camera detector that together provide a sensitivity enhancement of ~104. These studies indicate that the estrogen receptor in breast cancer cells can be visualized by fluorescence techniques, provided that the visualizing ligands have adequate affinity and specificity for the receptor and appropriate fluorescence characteristics, and provided that the fluorescence instrument has adequate sensitivity to observe fluorescence emission from cells treated with nM concentrations of the fluorescent agents.

Original languageEnglish (US)
Pages (from-to)4956-4965
Number of pages10
JournalCancer Research
Volume43
Issue number10
StatePublished - Oct 1 1983
Externally publishedYes

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

Fingerprint

Dive into the research topics of 'New Approach for Visualizing Estrogen Receptors in Target Cells Using Inherently Fluorescent Ligands and Image Intensification'. Together they form a unique fingerprint.

Cite this