Western blot analysis of extracts from various cell lines using Cadherin-17 Antibody (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower). As expected, Cadherin-17 protein is not detected in PANC-1 cells.Learn more about how we get our images.
Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing Myc/DDK-tagged full-length human CDH17 protein (hCDH17-Myc/DDK; +), using Cadherin-17 Antibody (upper), DYKDDDDK Tag Antibody #2368 (middle), and β-Actin (D6A8) Rabbit mAb #8457 (lower).Learn more about how we get our images.
For western blots, incubate membrane with diluted primary antibody in 5% w/v nonfat dry milk, 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.
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 June 2016
Protocol Id: 263
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.
Cadherin-17 Antibody recognizes endogenous levels of total Cadherin-17 protein. Based upon sequence alignment, this antibody is not predicted to cross-react with Cadherin-16 protein.
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues near the carboxy terminus of human Cadherin-17 protein. Antibodies are purified by protein A and peptide affinity chromatography.
Cadherins are a superfamily of transmembrane glycoproteins that contain cadherin repeats of approximately 100 residues in their extracellular domain. Cadherins mediate calcium-dependent cell-cell adhesion and play critical roles in normal tissue development (1). The classic cadherin subfamily includes N-, P-, R-, B-, and E-cadherins, as well as about ten other members that are found in adherens junctions, a cellular structure near the apical surface of polarized epithelial cells. The cytoplasmic domain of classical cadherins interacts with β-catenin, γ-catenin (also called plakoglobin), and p120 catenin. β-catenin and γ-catenin associate with α-catenin, which links the cadherin-catenin complex to the actin cytoskeleton (1,2). While β- and γ-catenin play structural roles in the junctional complex, p120 regulates cadherin adhesive activity and trafficking (1-4). Investigators consider E-cadherin an active suppressor of invasion and growth of many epithelial cancers (1-3). Research studies indicate that cancer cells have upregulated N-cadherin in addition to loss of E-cadherin. This change in cadherin expression is called the "cadherin switch." N-cadherin cooperates with the FGF receptor, leading to overexpression of MMP-9 and cellular invasion (3). Research studies have shown that in endothelial cells, VE-cadherin signaling, expression, and localization correlate with vascular permeability and tumor angiogenesis (5,6). Investigators have also demonstrated that expression of P-cadherin, which is normally present in epithelial cells, is also altered in ovarian and other human cancers (7,8).
Cadherin-17/Liver-Intestine-cadherin (CDH17/LI-cadherin) is a type-I transmembrane glycoprotein that belongs to the 7D-cadherin superfamily. Unlike classical cadherins, CDH17 is characterized by an extracellular domain with seven cadherin repeats and a short cytoplasmic domain that does not display any homology to the cytoplasmic domain of classical cadherins (9). CDH17 is a calcium-dependent homotypic cell-cell adhesion molecule that is selectively expressed on the basolateral surface of polarized epithelia lining the small intestine and colon of humans under normal physiological conditions (10). Research studies have demonstrated that CDH17 is aberrantly overexpressed in human gastric cancer and may serve as a novel oncogenic biomarker for this disease (11,12). At the molecular level, research studies have suggested that CDH17 exerts its oncogenicity in the context of gastric cancer through its ability to engage both NF-κB and MAPK signaling cascades (13,14).
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|42919S||100 µl||$ 255.0|