|OX40 Ligand/TNFSF4 in Mouse Splenocytes. OX40 Ligand/TNFSF4 was detected in immersion fixed mouse splenocytes using Goat Anti-Mouse OX40 Ligand/TNFSF4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1236) at 15 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; Catalog # NL001) and counterstained with DAPI (blue). Specific staining was localized to the plasma membrane and cytoplasm. View our protocol for Fluorescent ICC Staining of Non-adherent Cells.|
|IL‑2 Secretion Induced by OX40 Ligand/TNFSF4 and Neutralization by Mouse OX40 Ligand/TNFSF4 Antibody. In the presence of a cross-linking antibody, Mouse polyHistidine Monoclonal Antibody (10 µg/mL, Catalog # MAB050) and sub-optimal amounts of Mouse CD3 epsilon Monoclonal Antibody (Catalog # MAB484), Recombinant Mouse OX40 Ligand/TNFSF4 (Catalog # 1236-OX) stimulates IL‑2 secretion in mouse T cells in a dose-dependent manner (orange line), as measured by the Mouse IL‑2 Quantikine ELISA Kit (Catalog # M2000). Under these conditions, IL‑2 secretion elicited by Recombinant Mouse OX40 Ligand/TNFSF4 (30 ng/mL) is neutralized (green line) by increasing concentrations of Mouse OX40 Ligand/TNFSF4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1236). The ND50 is typically 0.2-0.6 µg/mL.|
OX40 Ligand (OX40L), also known as gp34, is a type II transmembrane glycoprotein belonging to the TNF superfamily. Murine OX40L cDNA encodes a 198 amino acid (aa) residue protein comprised of a 28 aa N-terminal cytoplasmic domain, a 20 aa transmembrane segment, and a 150 aa C-terminal extracellular domain (1). Human and murine OX40L share 46% sequence identity at the amino acid level (1). The OX40L is expressed on activated antigen presenting cells such as B cells, macrophages, dendritic cells, and on endothelial cells at the site of inflammation. The receptor for OX40L is OX40 (CD134) that is expressed predominantly on activated CD4+ T cells. Expression of OX40 is transient following engagement of T cell receptors (2). Ligation of OX40L by OX40 stimulates proliferation and differentiation of activated B cells, and increases immunoglobulin secretion (3, 4). The expression of OX40L on B cells is up-regulated by CD40 ligation (3). Engagement of the OX40-OX40L system has co-stimulatory effects on T cells by stimulating the production of cytokines by T helper cells and increasing the survival of memory T cells (2, 5). Blocking of the OX40-OX40L interaction in vitro inhibits co-stimulation resulting in decreased T cell proliferation and adhesion of T cells to endothelial cells. Inhibition of the OX40-OX40L interaction in disease models has beneficial effects in acute graft-versus-host disease, inflammatory bowel disease, and decreases the development of collagen-induced arthritis and experimental leishmaniasis (6).
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