Immunological detection of the oestradiol receptor protein in cell lines derived from the lymphatic system and the haematopoietic system: variability of specific hormone binding in vitro

J Endocrinol. 1992 Sep;134(3):397-404. doi: 10.1677/joe.0.1340397.

Abstract

Extracts of human MCF 7 mammary carcinoma cells, the human lymphoblastoid cell lines AEH 1 and IM 9, T-cell derived CCRF cells, HL 60 myeloic leukaemia cells and murine myeloma cells SP 0 and NS I were analysed for immunoreactivity with polyclonal goat antibodies raised against homogeneous preparations of C-terminal fragments (32 kDa) of porcine uterine oestradiol receptor (ER). Whole cells and low speed cytosols were analysed for specific oestradiol-binding activity. ERs were enriched from cell extracts by either fractionated ethanol precipitation (0-25% (v/v) ethanol) and/or microscale-immunoaffinity chromatography. Immunoreactive proteins of identical molecular weight (approximately 65 kDa) were detected in all cell lines examined. Whole cell binding assays showed specific oestradiol-binding activity in MCF 7, IM 9 and CCRF cells. Borderline binding was found in HL 60 myeloid cells. No specific binding could be detected in AEH 1, NS I and SP 0 cells. Identical results were obtained using agar-electrophoresis after dextran-coated charcoal treatment. Immunoaffinity purified ERs from MCF 7, AEH 1 and HL 60 cells were subjected to limited proteolysis, where identical tryptic fragments were generated. In conclusion, we have confirmed by immunological methods that ERs are expressed in a variety of cell lines derived from the immune system and the haematopoietic system. The lack of specific hormone binding or very low-affinity hormone binding in some of the cells examined may be due to post-translational events or point mutations.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Blotting, Western
  • Cell Line
  • Chromatography, Affinity / methods
  • Cytosol / chemistry
  • Electrophoresis, Agar Gel
  • Estradiol / metabolism
  • Hematopoietic System / chemistry*
  • Humans
  • Immune Sera
  • Immunophenotyping / methods
  • Lymphatic System / chemistry*
  • Protein Binding / physiology
  • Receptors, Estradiol / analysis*
  • Receptors, Estradiol / immunology
  • Receptors, Estradiol / metabolism
  • Tumor Cells, Cultured / metabolism

Substances

  • Immune Sera
  • Receptors, Estradiol
  • Estradiol