AffinityImmuno Products

Introduction

CX3CL1, commonly called fractalkine, is the sole human chemokine of the CX3C class and is unusual in existing both as a membrane-tethered molecule with a mucin-like stalk and, after proteolytic shedding, as a soluble mediator. Endothelial cells display the membrane-bound form following exposure to inflammatory signals such as interferon-gamma and tumor necrosis factor-alpha, allowing it to grab and firmly anchor circulating monocytes, T cells, and NK cells at the vessel wall. Once cleaved free, the soluble fragment instead acts as a classic chemoattractant, drawing monocytes and T lymphocytes toward sites of injury or inflammation through its single receptor, CX3CR1, which is concentrated on cytotoxic lymphocytes and macrophages. This chemokine-receptor pairing has been implicated in a wide array of conditions, including atherosclerosis, vasculitis, several cancers, and chronic inflammatory and neurological disorders.

Because fractalkine exists both as a vessel-wall adhesion molecule and, once shed, as a soluble monocyte/T-cell chemoattractant acting through CX3CR1, this assay supports research spanning vascular inflammation (atherosclerosis, vasculitis) and CX3CR1-driven neuroinflammatory and oncology studies.

Principle of the Assay

This kit uses a sandwich enzyme immunoassay format to detect Human CX3CL1. The microtiter plate wells are pre-coated with a CX3CL1-specific capture antibody, which binds the target protein present in standards or samples. A biotin-conjugated detection antibody, also specific to Human CX3CL1, is then added and binds a second epitope on the captured analyte. Avidin conjugated to horseradish peroxidase (HRP) is subsequently introduced and binds the biotin label. Upon addition of TMB substrate solution, a colorimetric signal develops only in wells where CX3CL1, the biotin-conjugated detection antibody, and the avidin-HRP conjugate are all present. The enzymatic reaction is stopped by the addition of sulphuric acid solution, and absorbance is measured at 450 nm (± 10 nm). CX3CL1 concentrations in unknown samples are determined by comparing their optical density values against a standard curve.