AffinityImmuno Products

Introduction

Interleukin-34 is a cytokine that binds and activates the receptor tyrosine kinase CSF1R, the same receptor used by colony-stimulating factor 1, despite sharing little sequence similarity with that ligand. Through CSF1R engagement it triggers ERK1/ERK2 phosphorylation and downstream signaling that drives the proliferation, survival, and differentiation of monocytes and macrophages. In tissues where CSF1R is the dominant driver of local myeloid development, such as the skin and central nervous system, IL-34 is required for generating and maintaining specialized resident macrophage populations, namely Langerhans cells and microglia, that arise from early embryonic precursors. IL-34 also promotes osteoclast differentiation and bone resorption and stimulates release of pro-inflammatory chemokines, giving it roles spanning innate immune signaling, tissue-resident macrophage biology, and skeletal remodeling.

Because IL-34 signals through the same CSF1R receptor targeted by the approved therapy pexidartinib and the investigational agent vimseltinib for tenosynovial giant cell tumor, this assay is relevant to macrophage-targeted drug development in addition to its established role maintaining microglia and Langerhans cell populations and driving osteoclast-mediated bone resorption.

Principle of the Assay

This kit uses a sandwich enzyme immunoassay format in which the microtiter plate wells are pre-coated with a capture antibody specific to Human IL-34. Standards or samples are pipetted into the appropriate wells along with a biotin-conjugated detection antibody that recognizes a distinct epitope on Human IL-34. Avidin conjugated to horseradish peroxidase (HRP) is then added to each well and allowed to incubate, linking enzymatic activity to the captured analyte complex. Upon addition of TMB substrate solution, a colorimetric signal develops exclusively in wells containing Human IL-34, the biotin-conjugated antibody, and avidin-HRP. The enzymatic reaction is stopped by the addition of sulphuric acid solution, and the resulting color change is measured spectrophotometrically at 450 nm (± 10 nm). Sample concentrations are determined by comparing the optical density of each sample against a standard curve generated from the kit standards.