EphB2 Antibody [Unconjugated]

Simple Western lane view shows lysates of COLO 205 human colorectal adenocarcinoma cell line and MBA‑MB‑468 human breast cancer cell line, loaded at 0.2 mg/mL. A specific band was detected for EphB2 at approximately 139 and 146 kDa (as indicated) using 20 µg/mL of Goat Anti-Human/Mouse EphB2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF467) followed by 1:50 dilution of HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF109). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.

Western blot shows 25 ng of Recombinant Human EphB2 Fc Chimera (Catalog # 5189-B2) and Recombinant Mouse EphB2 Fc Chimera (Catalog # 467-B2). PVDF Membrane was probed with 0.1 µg/mL of Goat Anti-Human/Mouse EphB2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF467) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF109). A specific band was detected for EphB2 at approximately 120 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 3.

EphB2 was detected in immersion fixed MBA‑MB‑468 human breast cancer cell line using Goat Anti-Human/Mouse EphB2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF467) at 5 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; Catalog # NL001) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

EphB2 was detected in immersion fixed frozen sections of embryonic mouse brain (15 d.p.c.) using 15 µg/mL Goat Anti-Mouse EphB2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF467) overnight at 4 °C. Tissue was stained with the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; Catalog # NL001) and counterstained (green). View our protocol for Fluorescent IHC Staining of Frozen Tissue Sections.

COLO 205 human colorectal adenocarcinoma cell line was stained with Goat Anti-Human/Mouse EphB2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF467, filled histogram) or isotype control antibody (Catalog #AB-108-C, open histogram), followed by Phycoerythrin-conjugated Anti-Goat IgG Secondary Antibody (Catalog # F0107). View our protocol for Staining Membrane-associated Proteins.

D3 mouse embryonic stem cell line was stained with Goat Anti-Human/Mouse EphB2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF467, filled histogram) or isotype control antibody (Catalog #AB-108-C, open histogram), followed by Phycoerythrin-conjugated Anti-Goat IgG Secondary Antibody (Catalog # F0107). View our protocol for Staining Membrane-associated Proteins.

EphB2 was detected in immersion fixed paraffin-embedded sections of human esophageal squamous cell carcinoma using Goat Anti-Human/Mouse EphB2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF467) at 3 µg/mL for 1 hour at room temperature followed by incubation with the Anti-Goat IgG VisUCyte™ HRP Polymer Antibody (Catalog # VC004). Tissue was stained using DAB (brown) and counterstained with hematoxylin (blue). Specific staining was localized to cytoplasm in cancer cells. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

EphB2 was detected in immersion fixed frozen sections of mouse embryo (13 d.p.c.) using Goat Anti-Human/Mouse EphB2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF467) at 1.7 µg/mL for 1 hour at room temperature followed by incubation with the Anti-Goat IgG VisUCyte™ HRP Polymer Antibody (Catalog # VC004). Tissue was stained using DAB (brown) and counterstained with hematoxylin (blue). Specific staining was localized to developing brain. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

• Reactivity: Hu, Mu
• Applications: WB, Simple Western, Flow, IHC, CyTOF-ready, ICC/IF
• Clonality: Polyclonal
• Host: Goat
• Conjugate: Unconjugated
• Concentration: LYOPH

EphB2 Antibody [Unconjugated] Summary

• Immunogen: Mouse myeloma cell line NS0-derived recombinant mouse EphB2
  Val27-Lys548
  Accession # P54763
• Specificity: Detects mouse and human EphB2 in direct ELISAs and Western blots. In Western blots, approximately 5% cross-reactivity with recombinant rat (rr) EphB1, recombinant mouse (rm) EphA8, rmEphA6, rmEphB6, rmEphA3, rmEphA4, rmEphA7, rrEphA5, recombinant human EphA1, rmEphA2 and rmEphB3 is observed.
• Details of Functionality: ActivityAssaywCitation not found None None None None None
• Source: N/A
• Isotype: IgG
• Clonality: Polyclonal
• Host: Goat
• Gene: EPHB2

Applications/Dilutions

• Dilutions:
  • Western Blot 0.1 ug/mL
  • Simple Western 20 ug/mL
  • Flow Cytometry 0.25 ug/10^6 cells
  • Immunohistochemistry 1-15 ug/mL
  • CyTOF-ready
  • Immunocytochemistry 5-15 ug/mL

Packaging, Storage & Formulations

• Storage: Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 6 months, -20 to -70 °C under sterile conditions after reconstitution.
• Buffer: Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied as a 0.2 µm filtered solution in PBS.
• Preservative: No Preservative
• Concentration: LYOPH
• Reconstitution Instructions: Reconstitute at 0.2 mg/mL in sterile PBS.

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for EphB2 Antibody [Unconjugated]

• CAPB
• Cek5
• Drt
• DRTEphB2
• EC 2.7.10
• EC 2.7.10.1
• EK5
• elk-related tyrosine kinase
• EPH receptor B2
• eph tyrosine kinase 3
• EphB2
• EPH-like kinase 5
• ephrin type-B receptor 2
• EPHT3MGC87492
• EPTH3
• Erk
• ERKHek5
• Hek5
• Nuk
• PCBC
• protein-tyrosine kinase HEK5
• Qek2
• Renal carcinoma antigen NY-REN-47
• Sek3
• Tyro5
• Tyrosine-protein kinase receptor EPH-3
• Tyrosine-protein kinase TYRO5

Background

EphB2, also known as Cek5, Nuk, Erk, Qek2, Tyro5, Sek3, Hek5, and Drt (1), is a member of the Eph receptor family which binds members of the ephrin ligand family. There are two classes of receptors, designated A and B. Both the A and B class receptors have an extracellular region consisting of a globular domain, a cysteine-rich domain, and two fibronectin type III domains. This is followed by the transmembrane region and the cytoplasmic region. The cytoplasmic region contains a juxtamembrane motif with two tyrosine residues which are the major autophosphorylation sites, a kinase domain, and a conserved sterile alpha motif (SAM) in the carboxy tail which contains one conserved tyrosine residue. Activation of kinase activity occurs after ligand recognition and binding. EphB2 has been shown to bind ephrin-B1, ephrin-B2, and ephrin-B3 (2, 3). The extracellular domains of human and mouse EphB2 share 99% amino acid identity. Only membrane-bound or
Fc‑clustered ligands are capable of activating the receptor in vitro. Soluble monomeric ligands bind the receptor but do not induce receptor autophosphorylation and activation (2). In vivo, the ligands and receptors display reciprocal expression (3). It has been found that nearly all the receptors and ligands are expressed in developing and adult neural tissue (3). The ephrin/Eph families also appear to play a role in angiogenesis (3).

1. Eph Nomenclature Committee [letter] (1997) Cell 90:403.
2. Flanagan, J.G. and P. Vanderhaeghen (1998) Annu. Rev. Neurosci. 21:309.
3. Pasquale, E.B. (1997) Curr. Opin. Cell Biol. 9:608.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Publications for EphB2 Antibody (AF467)(33)

Publications using AF467

• A Kischel, C Audouard, MA Fawal, A Davy Ephrin-B2 paces neuronal production in the developing neocortex BMC Dev. Biol., 2020;20(1):12. 2020 [PMID: 32404061] (In Vivo, Mouse)
• ST Wei, YC Huang, ML Hsieh, YJ Lin, WC Shyu, HC Chen, CH Hsieh Atypical chemokine receptor ACKR3/CXCR7 controls postnatal vasculogenesis and arterial specification by mesenchymal stem cells via Notch signaling Cell Death Dis, 2020;11(5):307. 2020 [PMID: 32366833] (Western Blot, Human)
• D Vreeken, CS Bruikman, SML Cox, H Zhang, R Lalai, A Koudijs, AJ van Zonnev, GK Hovingh, JM van Gils EPH receptor B2 stimulates human monocyte adhesion and migration independently of its EphrinB ligands J. Leukoc. Biol., 2020;0(0):. 2020 [PMID: 32337793] (IHC, Human)
• L Farr, S Ghosh, N Jiang, K Watanabe, M Parlak, R Bucala, S Moonah CD74 signaling links inflammation to intestinal epithelial cell regeneration and promotes mucosal healing Cell Mol Gastroenterol Hepatol, 2020;0(0):. 2020 [PMID: 32004754] (IHC, Mouse)
• YL Liu, AM Horning, B Lieberman, M Kim, CK Lin, CN Hung, CW Chou, CM Wang, CL Lin, NB Kirma, MA Liss, R Vasisht, EP Perillo, K Blocher, H Horng, JA Taverna, J Ruan, TE Yankeelov, AK Dunn, TH Huang, HC Yeh, CL Chen Spatial EGFR Dynamics and Metastatic Phenotypes Modulated by Upregulated EphB2 and Src Pathways in Advanced Prostate Cancer Cancers (Basel), 2019;11(12):. 2019 [PMID: 31805710] (Western Blot, Human)
• Z Zhang, M Ye, Q Li, Y You, H Yu, Y Ma, L Mei, X Sun, L Wang, W Yue, R Li, J Li, D Zhang The Schizophrenia Susceptibility Gene OPCML Regulates Spine Maturation and Cognitive Behaviors through Eph-Cofilin Signaling Cell Rep, 2019;29(1):49-61.e7. 2019 [PMID: 31577955] (Immunoprecipitation, Mouse)
• AS Ernst, LI Böhler, AM Hagenston, A Hoffmann, S Heiland, C Sticht, M Bendszus, M Hecker, H Bading, HH Marti, T Korff, R Kunze EphB2-dependent signaling promotes neuronal excitotoxicity and inflammation in the acute phase of ischemic stroke Acta Neuropathol Commun, 2019;7(1):15. 2019 [PMID: 30722785] (Simple Western, Mouse)
• PN Mimche, CM Lee, SM Mimche, M Thapa, A Grakoui, M Henkemeyer, TJ Lamb EphB2 receptor tyrosine kinase promotes hepatic fibrogenesis in mice via activation of hepatic stellate cells Sci Rep, 2018;8(1):2532. 2018 [PMID: 29416088] (ICC, Mouse)
• S Miura, A Suzuki Generation of Mouse and Human Organoid-Forming Intestinal Progenitor Cells by Direct Lineage Reprogramming Cell Stem Cell, 2017;0(0):. 2017 [PMID: 28943029] (IHC, Mouse)
• YH Liu, JW Tsai, JL Chen, WS Yang, PC Chang, PL Cheng, DL Turner, Y Yanagawa, TW Wang, JY Yu Ascl1 promotes tangential migration and confines migratory routes by induction of Ephb2 in the telencephalon Sci Rep, 2017;7(0):42895. 2017 [PMID: 28276447] (ICC, Mouse)
• N Tüysüz, L van Bloois, S van den Br, H Begthel, MM Verstegen, LJ Cruz, L Hui, LJ van der La, J de Jonge, R Vries, E Braakman, E Mastrobatt, JJ Cornelisse, H Clevers, D Ten Berge Lipid-mediated Wnt protein stabilization enables serum-free culture of human organ stem cells Nat Commun, 2017;8(0):14578. 2017 [PMID: 28262686] (IHC, Human)
• Can Gao Blocking the Interaction between EphB2 and ADDLs by a Small Peptide Rescues Impaired Synaptic Plasticity and Memory Deficits in a Mouse Model of Alzheimer's Disease J. Neurosci., 2016;36(47):11959-11973. 2016 [PMID: 27881781] (Western Blot, Rat)
• Induction of endoplasmic reticulum stress by deletion of Grp78 depletes Apc mutant intestinal epithelial stem cells Oncogene, 2016;0(0):. 2016 [PMID: 27819675] (IHC-P, Mouse)
• AK O'Neill, AA Kindberg, TK Niethamer, AR Larson, HH Ho, ME Greenberg, JO Bush Unidirectional Eph/ephrin signaling creates a cortical actomyosin differential to drive cell segregation J. Cell Biol., 2016;215(2):217-229. 2016 [PMID: 27810913] (IHC, Mouse)
• The transcription factor XBP1s restores hippocampal synaptic plasticity and memory by control of the Kalirin-7 pathway in Alzheimer model Mol Psychiatry, 2016;0(0):. 2016 [PMID: 27646263] (IHC-Fr, Mouse)
• EphrinB2 repression through ZEB2 mediates tumour invasion and anti-angiogenic resistance Nat Commun, 2016;7(0):12329. 2016 [PMID: 27470974] (Western Blot, Mouse)
• Feng Y, Sakamoto N, Wu R, Liu J, Wiese A, Green M, Green M, Akyol A, Roy B, Zhai Y, Cho K, Fearon E Tissue-Specific Effects of Reduced beta-catenin Expression on Adenomatous Polyposis Coli Mutation-Instigated Tumorigenesis in Mouse Colon and Ovarian Epithelium. PLoS Genet, 2015;11(11):e1005638. 2015 [PMID: 26528816] (IHC-P, Mouse)
• Dong L, Gao F, Wang X, Miao Y, Wang S, Wu Y, Li F, Wu J, Cheng X, Sun X, Yang X, Wang Z GluA2 trafficking is involved in apoptosis of retinal ganglion cells induced by activation of EphB/EphrinB reverse signaling in a rat chronic ocular hypertension model. J Neurosci, 2015;35(13):5409-21. 2015 [PMID: 25834064] (Western Blot, Rat)
• Aharon, Rotem, Janes, Peter W, Burgess, Anthony, Hamza, Kais, Klebaner, Fima, Lackmann, Martin A mathematical model for eph/ephrin-directed segregation of intermingled cells. PLoS ONE, 2014;9(12):e111803. 2014 [PMID: 25436892] (Western Blot, Human)
• Boyer-Di Ponio J, El-Ayoubi F, Glacial F, Ganeshamoorthy K, Driancourt C, Godet M, Perriere N, Guillevic O, Couraud P, Uzan G Instruction of circulating endothelial progenitors in vitro towards specialized blood-brain barrier and arterial phenotypes. PLoS ONE, 2014;9(1):e84179. 2014 [PMID: 24392113] (ICC, Human)
• Bouche E, Romero-Ortega M, Henkemeyer M, Catchpole T, Leemhuis J, Frotscher M, May P, Herz J, Bock H Reelin induces EphB activation. Cell Res, 2013;23(4):473-90. 2013 [PMID: 23318582]
• Attwood BK, Bourgognon JM, Patel S, Mucha M, Schiavon E, Skrzypiec AE, Young KW, Shiosaka S, Korostynski M, Piechota M, Przewlocki R, Pawlak R Neuropsin cleaves EphB2 in the amygdala to control anxiety. Nature, 2011;473(7347):372-5. 2011 [PMID: 21508957] (Neutralization, Mouse)
• Furukawa K, Sato T, Katsuno T, Nakagawa T, Noguchi Y, Tokumasa A, Yokote K, Yokosuka O, Saito Y Smad3 contributes to positioning of proliferating cells in colonic crypts by inducing EphB receptor protein expression. Biochem. Biophys. Res. Commun., 2011;405(4):521-6. 2011 [PMID: 21276420] (Western Blot, Mouse)
• Bush JO, Soriano P Ephrin-B1 regulates axon guidance by reverse signaling through a PDZ-dependent mechanism. Genes Dev., 2009;23(13):1586-99. 2009 [PMID: 19515977] (Immunoprecipitation, Mouse)
• Allan EH, Hausler KD, Wei T, Gooi JH, Quinn JM, Crimeen-Irwin B, Pompolo S, Sims NA, Gillespie MT, Onyia JE, Martin TJ EphrinB2 regulation by PTH and PTHrP revealed by molecular profiling in differentiating osteoblasts. J. Bone Miner. Res., 2008;23(8):1170-81. 2008 [PMID: 18627264] (Western Blot, Mouse)
• Lim BK, Matsuda N, Poo MM Ephrin-B reverse signaling promotes structural and functional synaptic maturation in vivo. Nat. Neurosci., 2008;11(2):160-9. 2008 [PMID: 18193042] (Immunoprecipitation, Xenopus)
• Tanaka M, Kamata R, Takigahira M, Yanagihara K, Sakai R Phosphorylation of ephrin-B1 regulates dissemination of gastric scirrhous carcinoma. Am. J. Pathol., 2007;171(1):68-78. 2007 [PMID: 17591954] (Western Blot, Mouse)
• Saraswathi V, Gao L, Morrow JD, Chait A, Niswender KD, Hasty AH Fish oil increases cholesterol storage in white adipose tissue with concomitant decreases in inflammation, hepatic steatosis, and atherosclerosis in mice. J. Nutr., 2007;137(7):1776-82. 2007 [PMID: 17585030] (IHC-P, Human)
• Chaudhari A, Mahfouz M, Fialho AM, Yamada T, Granja AT, Zhu Y, Hashimoto W, Schlarb-Ridley B, Cho W, Das Gupta TK, Chakrabarty AM Cupredoxin-cancer interrelationship: azurin binding with EphB2, interference in EphB2 tyrosine phosphorylation, and inhibition of cancer growth. Biochemistry, 2007;46(7):1799-810. 2007 [PMID: 17249693] (Immunoprecipitation, Human)
• Holmberg J, Genander M, Halford MM, Anneren C, Sondell M, Chumley MJ, Silvany RE, Henkemeyer M, Frisen J EphB receptors coordinate migration and proliferation in the intestinal stem cell niche. Cell, 2006;125(6):1151-63. 2006 [PMID: 16777604] (Immunoprecipitation, Mouse)
• Ogawa K, Wada H, Okada N, Harada I, Nakajima T, Pasquale EB, Tsuyama S EphB2 and ephrin-B1 expressed in the adult kidney regulate the cytoarchitecture of medullary tubule cells through Rho family GTPases. J. Cell. Sci., 2006;119(0):559-70. 2006 [PMID: 16443753] (IHC, Mouse)
• Nakada M, Niska JA, Tran NL, McDonough WS, Berens ME EphB2/R-Ras signaling regulates glioma cell adhesion, growth, and invasion. Am. J. Pathol., 2005;167(2):565-76. 2005 [PMID: 16049340] (Western Blot, Human)
• Nakada M, Niska JA, Miyamori H, McDonough WS, Wu J, Sato H, Berens ME The phosphorylation of EphB2 receptor regulates migration and invasion of human glioma cells. Cancer Res., 2004;64(9):3179-85. 2004 [PMID: 15126357] (Immunoprecipitation, Human)

Reviews for EphB2 Antibody (AF467) (5)

Average Rating: 4.2

(Based on 5 reviews)

We have 5 reviews tested in 2 species: Mouse, Other.

reviewed by: STAVROULA THEOFANOUS WB Mouse 07/05/2018

Summary

• Application: Western Blot
• Sample Tested: brain and spinal cord
• Species: Mouse
• Lot: CFK071804A

Comments

• Comments: Blot Loading
  • ANTIBODY: Human/Mouse EphB2Antibody cat noAF542
  Lane1: Precision Plus Protein™ Dual Color Standards7ul
  Lane2:Ephb1 receptor (SC tissue from naive C57BL6j mouse)
  Western blot experiment details
  Gel:4-20% SDS page
  Tranfer apparatus: Trans-Blot® Turbo™ Transfer System
  Membrane: midi size nitrocellulose
  Time: 25min at 75-125V
  Primary ab:Goat –rat EphB1 Ab 1mg for 20h
  Secondary ab: rabbit a-goat 1:5,000 for 1h
  Substrate: HRP for 15min

reviewed by: STAVROULA THEOFANOUS IP Mouse 04/16/2018

Summary

• Application: Immunoprecipitation
• Sample Tested: brain and spinal cord
• Species: Mouse
• Lot: CVT0311121

Comments

• Comments: Blot Loading
  • ANTIBODY: Human/Mouse EphB2Antibody cat noAF467
  Lane1: PreĐision Plus Protein™ Dual Color Standards8ul
  Lane2:Ephb2 receptors IP 1mg protein (SC tissue)
  Lane3:Ephb2 receptors no IP 25ug (SC tissue)
  Lane4:Goat GFP IP (negative control instead of goat IgG)1mg (SC tissue)
  Lane5:Ephb2 receptors IP 1mg (SC tissue)
  Lane6:Ephb2 receptors no IP 25ug (SC tissue)
  Lane7:Goat GFP IP (negative control instead of goat IgG)1mg (SC tissue)
  Western blot experiment details
  Gel:4-20% SDS page
  Tranfer apparatus: Trans-Blot® Turďo™ Transfer System
  Membrane: midi size nitrocellulose
  Time: 25min at 75-125V
  Primary ab:Goat –m/h EphB2 Ab 1mg for 20h
  Secondary ab: donkey a-goat 1:15,000 for 1h
  Substrate: HRP for 15min

reviewed by: Anonymous Simple Western Mouse 07/12/2017

Summary

• Application: Simple Western
• Sample Tested: mouse brain lysate
• Species: Mouse

Comments

• Comments: Simple Western: anti-EphB2 antibody [biotechne AF467]
  Simple Western lane view shows a specific 128 kDa signal for EphB2 in 0.5 mg/ml of mouse brain lysate (lysis buffer from ARY014 proteome profiler kit). This run was performed under reducing conditions using the 12-230 kDa separation system in combination with a secondary anti-goat-HRP antibody (biotechne, HAF109, 1:50).

reviewed by: Anonymous WB Mouse 01/08/2015

Summary

• Application: Western Blot
• Sample Tested: See PMID 21276420
• Species: Mouse

Blocking

• Blocking Details: See PMID 21276420

Details

• Detection Notes: See PMID 21276420

Comments

• Comments: Published: See PMID 21276420Independent review from antibodyresource.com

reviewed by: Anonymous WB Other 01/08/2015

Summary

• Application: Western Blot
• Sample Tested: See PMID 22144690
• Species: Other

Blocking

• Blocking Details: See PMID 22144690

Details

• Detection Notes: See PMID 22144690

Comments

• Comments: Published: See PMID 22144690Independent review from antibodyresource.com

Bioinformatics

• Gene Symbol: EPHB2
• Uniprot:
  • P54763()