Western blot analysis of extracts from HL-60, MKN-45 and HCC827 cells using DCBLD2 Antibody.
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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
DCBLD2 Antibody detects endogenous levels of total DCBLD2 protein.Species Reactivity:
Human, Mouse, RatSpecies predicted to react based on 100% sequence homology:
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues near the carboxy terminus of human DCBLD2 protein. Antibodies are purified by protein A and peptide affinity chromatography.
DCBLD2 (discoidin, CUB and LCCL domain-containing 2; also known as ESDN and CLCP1) is a type I transmembrane protein that is structurally similar to neuropilin family proteins and contains the longest known amino-terminal secretory signal sequence among eukaryotic proteins (1). Highly expressed in nerve bundles, vascular smooth muscle cells and upregulated following vascular injury, DCBLD2 may be involved in a wide range of functions in the nervous and vascular systems (1,2). Studies have found DCBLD2 to be upregulated in several types of lung cancer with an especially high frequency in metastatic lesions and lymph node metastasis, implicating a role in the process of tumor progression and metastatic capability (3). DCBLD2 has also been identified as part of a complex EGF phosphotyrosine signaling network, serving as a novel tyrosine phosphorylation target of EGF signaling in human cancer cells (4).
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