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The development of a CXCR2 chemokine receptorbinding assay using the LEADseeker Multimodality Imaging System

Key words: CXCR2 • GPCR • receptor binding assay • LEADseeker • SPA Imaging Beads


Chemo-attractant cytokines (chemokines) are low molecular weight, pro-inflammatory cytokines with potent leukocyte activation and/or chemotactic activity in vivo (1). Chemokines are classified into four families (CXC, CC, C, and CXC3) based on the number and sequential relationship of the first two of four conserved cysteine residues. They induce their biological effect by binding to chemokine receptors that belong to the superfamily of G protein-coupled seven-transmembrane domain receptors (2).

The CXC chemokines are pivotal mediators that activate or inhibit angiogenesis by interacting with cognate receptors expressed by endothelial cells (3). CXCR2 is a major chemokine receptor expressed in neutrophils and binds chemokines with a Glu-Leu-Arg (ELR) sequence in the amino terminus; IL-8, GRO-α, GRO-β, GRO-γ, NAP-2, ENA-78, and GCP-2 (4).

This application note describes a 384-well CXCR2 chemokine receptor-binding assay which has been developed using the LEADseeker™ Multimodality Imaging System.


Materials
Products used
LEADseeker Multimodality Imaging System 18-1140-71

Wheat Germ Agglutinin (WGA) PS SPA RPNQ0260
Imaging Beads

[125I]GRO-a IM305


Other materials required
Human recombinant CXCR2 receptor membrane preparation (Euroscreen ES-145-M)

Human recombinant IL-8 (Sigma 123K0714)

Human recombinant GRO-α(Sigma G0657)

Costar™ solid
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