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Enzyme-Linked Immunosorbent Assay01:33

Enzyme-Linked Immunosorbent Assay

In 1971, Peter Perlman and Eva Engvall developed an Enzyme-linked immunosorbent assay (ELISA or EIA). ELISA differs from western blot in that the assays are conducted in microtiter plates or in vivo rather than on an absorbent membrane.
There are many different types of ELISAs, but they all involve an antibody molecule whose constant region binds an enzyme, leaving the variable region free to bind its specific antigen.  Enzyme-substrate reaction allows the antigen to be visualized or quantified.

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Screening for Phytoestrogens using a Cell-based Estrogen Receptor β Reporter Assay
06:07

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Published on: June 7, 2020

A simple heterogeneous one-step assay for screening estrogenic compounds.

Tuomas Huovinen1, Kalle Rytkönen, Urpo Lamminmäki

  • 1Department of Biotechnology, University of Turku, Turku, Finland. tuomas.huovinen@utu.fi

Biotechnology Letters
|September 19, 2012
PubMed
Summary

A new bioassay detects estrogenic and anti-estrogenic compounds by measuring co-activator recruitment to estrogen receptor alpha (ERα). This simple assay aids in discovering therapeutics for hormonal diseases like breast cancer.

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Endocrinology

Background:

  • Estrogen receptor (ER) modulators pose health risks, yet ER antagonists are crucial for developing therapeutics against hormonal diseases, including breast cancer.
  • Screening for novel estrogenic and anti-estrogenic compounds is vital for advancing treatments for hormone-dependent conditions.

Purpose of the Study:

  • To develop a novel, simple bioassay for detecting estrogenic and anti-estrogenic compounds.
  • To utilize ligand-dependent co-activator recruitment for ERα activity assessment.

Main Methods:

  • The assay employs purified Renilla luciferase-tagged ERα and in vivo-biotinylated steroid receptor co-activator 1 (SRC-1).
  • Compounds are incubated with ERα and SRC-1 in streptavidin-coated microtiter wells, followed by a washing step and luminescence measurement.
  • The method avoids chemical labeling of assay components.

Main Results:

  • The bioassay demonstrates high sensitivity, detecting estradiol (E2) at concentrations as low as 25 pM.
  • The assay exhibits a wide dynamic range, spanning over four orders of magnitude.
  • Ligand-dependent recruitment of SRC-1 to ERα is effectively measured.

Conclusions:

  • This novel bioassay provides a simple and sensitive method for screening estrogenic and anti-estrogenic compounds.
  • The assay's capabilities support the search for new therapeutics targeting hormonal diseases.
  • The method's efficiency and broad dynamic range make it valuable for drug discovery efforts.