Detects canine TNF-alpha direct ELISAs and Western blots. In direct ELISAs, approximately 70% cross-reactivity with recombinant human TNF‑ alpha, 50% cross-reactivity with recombinant rhesus macaque TNF-alpha, 35% cross-reactivity with recombinant equine TNF‑ alpha, recombinant cotton rat TNF‑ alpha and recombinant rat TNF-alpha, 25% cross-reactivity with recombinant mouse TNF-alpha, and 10% cross-reactivity with recombinant porcine TNF‑ alpha is observed.
Polyclonal Goat IgG
E. coli-derived recombinant canine TNF-alpha Val77-Leu233 Accession # P51742
Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied as a 0.2 µm filtered solution in PBS.
<0.10 EU per 1 μg of the antibody by the LAL method.
Measured by its ability to neutralize TNF‑ alpha -induced cytotoxicity in the L‑929 mouse fibroblast cell line. Matthews, N. and M.L. Neale (1987) in Lymphokines and Interferons, A Practical Approach. Clemens, M.J. et al. (eds): IRL Press. 221. The Neutralization Dose (ND50) is typically 0.4-1.6 µg/mL in the presence of 100 ng/mL Recombinant Canine TNF‑ alpha and 1 µg/mL actinomycin D.
Please Note: Optimal dilutions should be determined by each laboratory for each application. General Protocols are available in the Technical Information section on our website.
Cytotoxicity Induced by TNF‑ alpha and Neutralization by Canine TNF‑ alpha Antibody.
Recombinant Canine TNF‑ alpha (Catalog # 1507‑CT) induces cytotoxicity in the the L‑929 mouse fibroblast cell line in a dose-dependent manner (orange line), as measured by crystal violet staining. Cytotoxicity elicited by Recombinant Canine TNF‑ alpha (100 ng/mL) is neutralized (green line) by increasing concentrations of Goat Anti-Canine TNF‑ alpha Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1507). The ND50 is typically 0.4-1.6 µg/mL in the presence of the metabolic inhibitor actinomycin D (1 µg/mL).
TNF‑ alpha in Canine PBMCs. TNF‑ alpha was detected in immersion fixed canine peripheral blood mononuclear cells (PBMCs) stimulated with PMA and calcium ionomycin using Goat Anti-Canine TNF‑ alpha Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1507) 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 cytoplasm. View our protocol for Fluorescent ICC Staining of Non-adherent Cells.
Preparation and Storage
Reconstitute at 0.2 mg/mL in sterile PBS.
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. *Small pack size (SP) is shipped with polar packs. Upon receipt, store it immediately at -20 to -70 °C
Stability & Storage
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
12 months from date of receipt, -20 to -70 °C as supplied.
1 month, 2 to 8 °C under sterile conditions after reconstitution.
6 months, -20 to -70 °C under sterile conditions after reconstitution.
Tumor necrosis factor alpha (TNF-alpha ), also known as cachectin and TNFSF2, is the prototypic ligand of the TNF superfamily. It is a pleiotropic molecule that plays a central role in inflammation, apoptosis, and immune system development. TNF-alpha is produced by a wide variety of immune and epithelial cell types (1, 2). Canine TNF-alpha consisits of a 35 amino acid (aa) cytoplasmic domain, a 21 aa transmembrane segment, and a 177 aa extracellular domain (ECD) (3). Within the ECD, canine TNF-alpha shares 84-94% aa sequence identity with equine, feline, human, porcine, and rhesus and 69-77% with bovine, cotton rat, mouse, and rat with TNF-alpha. The 26 kDa type 2 transmembrane protein is assembled intracellularly to form a noncovalently linked homotrimer (4). Ligation of this complex induces reverse signaling that promotes lymphocyte co-stimulation but diminishes monocyte responsiveness (5). Cleavage of membrane bound TNF-alpha by TACE/ADAM17 releases a 55 kDa soluble trimeric form of TNF-alpha (6, 7). TNF-alpha trimers bind the ubiquitous TNF RI and the hematopoietic cell-restricted TNF RII, both of which are also expressed as homotrimers (1, 8). TNF-alpha regulates lymphoid tissue development through control of apoptosis (2). It also promotes inflammatory responses by inducing the activation of vascular endothelial cells and macrophages (2). TNF-alpha is a key cytokine in the development of several inflammatory disorders (9). It contributes to the development of type 2 diabetes through its effects on insulin resistance and fatty acid metabolism (10, 11).
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