dep-1 Antibody
Description
DEP-1 Structure and Biological Roles
DEP-1 is a transmembrane receptor phosphatase composed of:
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Extracellular domain: Eight fibronectin type III repeats involved in cell-cell interactions .
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Transmembrane domain: Anchors the protein to the plasma membrane .
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Intracellular catalytic domain: Mediates dephosphorylation of tyrosine residues on target proteins like VEGFR2, PDGFR, EGFR, and Src kinase .
Key biological functions include:
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Tumor suppression: Dephosphorylation of growth factor receptors (e.g., EGFR, MET) to inhibit proliferation and promote contact inhibition .
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Endothelial regulation: Modulation of VEGF-dependent Src and Akt signaling to control angiogenesis and cell survival .
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Immune modulation: Attenuation of T-cell receptor signaling and immune synapse formation .
Available DEP-1 Antibodies
Commercial and research-grade DEP-1 antibodies vary in host species, applications, and target specificity:
Notes:
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WB: Western blot; IP: Immunoprecipitation; FCM: Flow cytometry; ICC/IF: Immunocytochemistry/immunofluorescence.
Western Blot Validation
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HeLa and Raji cell lysates show a specific ~220 kDa band for DEP-1 .
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siRNA-mediated DEP-1 knockdown reduces protein levels by >70% in THP-1 and Ln229 cells .
Functional Studies
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EGFR Regulation: DEP-1 knockdown increases EGFR phosphorylation (Y1173) in endosomes and enhances HeLa cell proliferation (p < 0.001) .
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VEGF Signaling: DEP-1 dephosphorylates VEGFR2 (Y1175) and promotes Src-dependent Akt activation, essential for endothelial survival .
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FLT3 Interaction: Proximity ligation assays confirm DEP-1 binds FLT3 in COS7 cells, modulating leukemic cell signaling .
Key Research Findings
Clinical and Therapeutic Relevance
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Cancer Biomarker: Reduced DEP-1 expression is linked to colon, thyroid, and breast cancers .
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Therapeutic Target: Antisense oligonucleotides targeting DEP-1 improve metabolic phenotypes in obesity models .
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Drug Resistance: DEP-1 overexpression in leukemia cells reduces FLT3 inhibitor efficacy .
Research Tools and Protocols
Q&A
DEP-1 (CD148) is a receptor-type protein tyrosine phosphatase critical for regulating cellular signaling pathways. Below are structured FAQs addressing key research considerations, methodologies, and challenges in DEP-1 antibody applications, synthesized from peer-reviewed studies and technical documentation .
Advanced Research Questions
How to resolve discrepancies in DEP-1 molecular weight across studies?
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Cause: Differential glycosylation (e.g., HeLa vs. Raji cells) .
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Method: Deglycosylate lysates with PNGase F and re-probe with anti-DEP-1 (Clone 261922) .
What experimental strategies validate DEP-1’s role in angiogenesis?
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In vitro: Use bivalent anti-DEP-1 antibodies (e.g., Ab1) to inhibit endothelial cell proliferation .
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In vivo: Corneal angiogenesis assays in knockout mice show impaired vascularization .
How to optimize DEP-1 immunoprecipitation for phosphoproteomics?
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Antibody: Use agarose-conjugated Clone 143-41 for high-affinity pull-downs .
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Lysis buffer: Include phosphatase inhibitors (e.g., sodium orthovanadate) to preserve phosphorylation states .
Methodological Challenges & Solutions
Addressing non-specific binding in flow cytometry with anti-DEP-1
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Issue: Cross-reactivity with fibronectin domains in serum proteins.
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Fix: Pre-clear lysates with Protein A/G beads and use Fc-blocking reagents .
Reconciling DEP-1’s dual role in tyrosine phosphorylation
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Context-dependent activity: While DEP-1 generally suppresses TCR signaling, it induces phosphorylation in tumor cells via serine/threonine kinases .
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Experimental design: Compare isogenic cell lines (wild-type vs. DEP-1 KO) under identical stimulation conditions .
