Intracellular Staining by Flow Cytometry
|Detection of IL‑1ra/IL‑1F3 in PBMCs by Flow Cytometry. Peripheral Blood Mononuclear Cells (PBMCs) either (A) untreated or (B) treated with LPS were stained with Mouse Anti-Human CD14 Fluorescein‑conjugated Monoclonal Antibody (Catalog # FAB3832F) and Mouse Anti-Human IL‑1ra/IL‑1F3 APC‑conjugated Monoclonal Antibody (Catalog # IC280A). Quadrant markers were set based on control antibody staining with Mouse IgG2A Allophycocyanin Isotype Control (Catalog # IC003A).To facilitate intracellular staining, cells were fixed with Flow Cytometry Fixation Buffer (Catalog # FC004) and permeabilized with Flow Cytometry Permeabilization/Wash Buffer I (Catalog # FC005). View our protocol for Staining Intracellular Molecules.|
IL-1ra was originally isolated from the urine of patients with monocytic leukemia and has also been purified from adherent monocytes. The naturally occurring, fully glycosylated form has an apparent molecular weight of about 25,000 Daltons. The protein shows 26% amino acid homology to IL-1 beta and 19% homology to IL-1 alpha. It will compete with either factor for receptor binding, but does not interact with either one. Human IL-1ra will bind to both types of IL-1 receptor (I and II) on human cells, but reportedly will not block binding to the type II receptor on murine pre-B cell lines. The recombinant, non-glycosylated form of IL-1ra blocks binding of IL-1 to its receptor equally as well as the naturally-occurring, glycosylated form. The IL-1ra has been shown to block the inflammatory responses induced by IL-1 both in vitro and in vivo.