The study of CXCR3 and CCR7 pharmacology using [35S]GTPgammaS exchange assays in cell membranes and permeabilized peripheral blood lymphocytes

J Immunol Methods. 2003 Feb;273(1-2):15-27. doi: 10.1016/s0022-1759(02)00415-5.

Abstract

The GTPgammaS exchange assay is a functional model corresponding to the first step of G protein-coupled receptor activation. We provide simple methodologies and controls for setting up GTPgammaS exchange assays in both cell membranes and permeabilized peripheral blood lymphocytes. Specifically, we use guanosine 5'-[35S]triphospate ([35S]GTPgammaS) exchange, in concert with radioligand binding assays, to assess the expression and function of two chemokine receptors important in the trafficking of T lymphocytes: CXCR3 and CCR7. The studies presented here illustrate the utility of GTPgammaS exchange assays in the study of chemokine receptor pharmacology.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding, Competitive
  • Cell Membrane / metabolism
  • Cell Membrane Permeability
  • Guanosine 5'-O-(3-Thiotriphosphate) / metabolism*
  • Guanosine Diphosphate / metabolism
  • Humans
  • Kinetics
  • Lymphocytes / metabolism*
  • Radioligand Assay / methods
  • Receptors, CCR7
  • Receptors, CXCR3
  • Receptors, Chemokine / metabolism*
  • Regression Analysis
  • Scintillation Counting
  • Tumor Cells, Cultured

Substances

  • CCR7 protein, human
  • CXCR3 protein, human
  • Receptors, CCR7
  • Receptors, CXCR3
  • Receptors, Chemokine
  • Guanosine Diphosphate
  • Guanosine 5'-O-(3-Thiotriphosphate)