High pressure liquid chromatographic detection of intracellular retinoid binding proteins from cultured cell and tumor cytosols

Elizabeth A. Allegretto, Michael A. Kelly, Carol A. Donaldson, Norman Levine, J. Wesley Pike, Mark R. Haussler

Research output: Contribution to journalArticle

12 Scopus citations

Abstract

We report the first application of high pressure liquid chromatography (HPLC) in the rapid detection of cellular retinoic acid binding protein (CRABP) and cellular retinol binding protein (CRBP). Cytosols from cultured cells (3T6 and MCF-7) or from tumors (melanoma and ovarian) were labeled with [3H]retinoic acid (30 Ci/mmol) and [3H]retinol (43 Ci/mmol) and analyzed via HPLC employing a 60 cm TSK 3000 sw column. In each case CRABP and CRBP were readily detectable at an elution volume of 22.5 ml, consistent with their molecular weights of 14,600. Identity of the binding protein peaks was established by saturability, specificity, and selective inhibition of binding by an organomercurial. Thus, this method, which resolves CRABP and CRBP in crude mixtures from the majority of cytosolic proteins, should be a valuable tool in the evaluation of vitamin A-binding protein interactions and their biological significance.

Original languageEnglish (US)
Pages (from-to)75-81
Number of pages7
JournalBiochemical and Biophysical Research Communications
Volume116
Issue number1
DOIs
StatePublished - Oct 14 1983

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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