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Flow Cytometric Analysis of CD324 (E-cadherin) on human breast adenocarcinoma (left) and human embryonic stem (ES) cells (right). Human breast adenocarcinoma cell line MCF-7 (ATCC, HTB-22) and H9 human ES Cells (WiCell, Madison, WI) were harvested with Cell Dissociation Buffer (Life Technologies). Please note that the epitope is sensitive to Accutase™ detachment. Cells were stained with either PE Mouse IgG1, k isotype control (Cat. No. 554680; dashed line) or PE Mouse anti-human CD324 antibody (Cat. No. 562870; solid line) at matched concentrations. Histograms were derived from gated events based on light scattering characteristics of MCF-7 or human ES cells. Flow cytometry was performed on a BD FACSCanto™ II flow cytometry system. Staining with this antibody can also performed on cells that have been fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655) and permeabilized with BD Phosflow™ Perm/Wash Buffer I (Cat. No. 557885). However, the brightness of the staining decreases after cell fixation and permeabilization.
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BD Pharmingen™ PE Mouse Anti-Human CD324 (E-Cadherin)
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Recommended Assay Procedures
In addition to standard cell surface staining of live cells, this antibody conjugate may also be used to stain cells that have been fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655) and permeabilized with BD Phosflow™ Perm/Wash Buffer I (Cat. No. 557885). Because the brightness of the staining decreases after cell fixation and permeabilization, the reagent titer may need to be adjusted.
Product Notices
- This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
- An isotype control should be used at the same concentration as the antibody of interest.
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- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
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The 67A4 monoclonal antibody specifically recognizes the extracellular domain of human E-Cadherin (CD324). E-Cadherin is a 120-kDa transmembrane glycoprotein that is localized in the adherens junctions of epithelial cells. There it interacts with the cytoskeleton through the associated cytoplasmic catenin proteins. In addition to being a calcium-dependent adhesion molecule, E-Cadherin is also a critical regulator of epithelial junction formation. Its association with catenins is necessary for cell-to-cell adhesion. These E-Cadherin/catenin complexes associate with cortical actin bundles at both the zonula adherens and the lateral adhesion plaques. Tyrosine phosphorylation can disrupt these complexes, leading to changes in cell adhesion properties. E-Cadherin expression is often down-regulated in highly invasive, poorly differentiated carcinomas. Increased expression of E-Cadherin in these cells reduces their invasiveness. Thus, loss of expression or function of E-Cadherin appears to be an important step in tumorigenic progression. Pluripotent stem cells express E-Cadherin. Upon differentiation, an epithelial to mesenchymal transition results in the loss of E-cadherin expression and a gain in the expression of N-cadherin.

Development References (5)
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Behrens J, Vakaet L, Friis R, et al. Loss of epithelial differentiation and gain of invasiveness correlates with tyrosine phosphorylation of the E-cadherin/beta-catenin complex in cells transformed with a temperature-sensitive v-SRC gene.. J Cell Biol. 1993; 120(3):757-66. (Biology). View Reference
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Bühring HJ, Müller T, Herbst R, et al. The adhesion molecule E-cadherin and a surface antigen recognized by the antibody 9C4 are selectively expressed on erythroid cells of defined maturational stages.. Leukemia. 1996; 10(1):106-16. (Clone-specific: Flow cytometry). View Reference
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Cepek KL, Shaw SK, Parker CM, et al. Adhesion between epithelial cells and T lymphocytes mediated by E-cadherin and the alpha E beta 7 integrin.. Nature. 1994; 372(6502):190-3. (Biology). View Reference
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D'Amour KA, Agulnick AD, Eliazer S, Kelly OG, Kroon E, Baetge EE. Efficient differentiation of human embryonic stem cells to definitive endoderm.. Nat Biotechnol. 2005; 23(12):1534-41. (Biology). View Reference
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Takeichi M. The cadherins: cell-cell adhesion molecules controlling animal morphogenesis.. Development. 1988; 102(4):639-55. (Biology). View Reference
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