The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 - 5 µg/ml. This antibody has only been tested in WB against the recombinant fragment used as immunogen. We have no data on the detection of endogenous protein.
Use a concentration of 3 µg/ml.
Use 1µg for 106 cells. (methanol fixed cells)
ab170192-Mouse monoclonal IgG2b, is suitable for use as an isotype control with this antibody.
Use at an assay dependent concentration.
FunctionCleaves the membrane-bound precursor of TNF-alpha to its mature soluble form. Responsible for the proteolytical release of soluble JAM3 from endothelial cells surface. Responsible for the proteolytic release of several other cell-surface proteins, including p75 TNF-receptor, interleukin 1 receptor type II, p55 TNF-receptor, transforming growth factor-alpha, L-selectin, growth hormone receptor, MUC1 and the amyloid precursor protein. Also involved in the activation of Notch pathway.
Tissue specificityUbiquitously expressed. Expressed at highest levels in adult heart, placenta, skeletal muscle, pancreas, spleen, thymus, prostate, testes, ovary and small intestine, and in fetal brain, lung, liver and kidney.
DomainMust be membrane anchored to cleave the different substrates. The cytoplasmic domain is not required for the this activity. Only the catalytic domain is essential to shed TNF and p75 TNFR. The conserved cysteine present in the cysteine-switch motif binds the catalytic zinc ion, thus inhibiting the enzyme. The dissociation of the cysteine from the zinc ion upon the activation-peptide release activates the enzyme.
Post-translational modificationsThe precursor is cleaved by a furin endopeptidase. Phosphorylated. Stimulation by growth factor or phorbol 12-myristate 13-acetate induces phosphorylation of Ser-819 but decreases phosphorylation of Ser-791.
ADAM17 antibody (ab57484) used in immunohistochemistry at 3ug/ml on formalin fixed and paraffin embedded human small Intestine.
Western blot - ADAM17 antibody (ab57484)
Western blot against tagged recombinant protein immunogen using ab57484 ADAM17 antibody at 1ug/ml. Predicted band size of immunogen is 37 kDa
Immunocytochemistry/ Immunofluorescence - ADAM17 antibody (ab57484)Image taken from an Abreview published in February 2008
ab57484 was used to detect mouse ADAM17 overexpressed in Hela cells. The transfected cells were incubated with ab57484 (10 µg/ml ) for 1 hour at 22°C. For further details please refer to the Abreview.
Flow Cytometry - ADAM17 antibody (ab57484)
Overlay histogram showing HeLa cells stained with ab57484 (red line). The cells were fixed with methanol (5 min) and incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab57484, 1µg/1x106 cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG2b [PLPV219] (ab91366, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a decreased signal in HeLa cells fixed with 4% paraformaldehyde (10 min) used under the same conditions.
Please note that Abcam do not have data for use of this antibody on non-fixed cells. We welcome any customer feedback.
Western blot - Anti-ADAM17 antibody (ab57484)
Anti-ADAM17 antibody (ab57484) at 1/1000 dilution + whole cell lysate prepared from MCF7 cells at 100000 cells
Secondary HRP conjugated sheep anti-mouse polyclonal at 1/2000 dilution developed using the ECL technique