Western blot analysis of extracts from serum-starved A-431 cells, untreated (-) or treated with Human Epidermal Growth Factor (hEGF) #8916 (100 ng/ml, 30 min) (+) using Phospho-DR6 (Ser562) Antibody (top) or α-Tubulin (11H10) Rabbit mAb #2125 (bottom).
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Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C. Do not aliquot the antibody.
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody datasheet or product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised November 2013
Reprobing of an existing membrane is a convenient means to immunoblot for multiple proteins independently when only a limited amount of sample is available. It should be noted that for the best possible results a fresh blot is always recommended. Reprobing can be a valuable method but with each reprobing of a blot there is potential for increased background signal. Additionally, it is recommended that you verify the removal of the first antibody complex prior to reprobing so that signal attributed to binding of the new antibody is not leftover signal from the first immunoblotting experiment. This can be done by re-exposing the blot to ECL reagents and making sure there is no signal prior to adding the next primary antibody.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.
posted June 2005
revised October 2016
Protocol Id: 10
Phospho-DR6 (Ser562) Antibody recognizes endogenous levels of DR6 protein only when phosphorylated at Ser562. This antibody may recognize DR6 with dual phosphorylation at Ser562 and Ser565.Species Reactivity:
Human, Mouse, Rat
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Ser562 of human DR6 protein. Antibodies are purified by protein A and peptide affinity chromatography.
The tumor necrosis factor receptor family, which includes TNF-RI, Fas, DR3, DR4, DR5, and DR6, plays an important role in the regulation of apoptosis in various physiological systems (1,2). The receptors are activated by a family of cytokines that include TNF, FasL, and TRAIL. They are characterized by a highly conserved extracellular region containing cysteine-rich repeats and a conserved intracellular region of about 80 amino acids termed the death domain (DD). The DD is important for transducing the death signal by recruiting other DD containing adaptor proteins (FADD, TRADD, RIP) to the death-inducing signaling complex (DISC), resulting in activation of caspases.
DR6, also known as TNFRSF21, is a TNFR family member able to induce apoptosis as well as activation of NF-κB and JNK (3). Expression of DR6 is upregulated by NF-κB signaling (4). DR6 appears to play a critical role in the activation and differentiation of T and B lymphocytes (5,6). In the nervous system β-amyloid precursor protein (APP) activates DR6 to trigger neuronal degeneration (7).
Phospho-DR6 (Ser562) Antibody is directed at a site that was identified at Cell Signaling Technology (CST) using PhosphoScan®, our proprietary LC-MS/MS platform for modification site discovery using an Akt substrate antibody (8). Please visit PhosphoSitePlus®, our modification site knowledgebase, at www.phosphosite.org for more information.
Explore pathways + proteins related to this product.
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