EphA2 Antibody [Unconjugated]

Western blot shows lysates of A431 human epithelial carcinoma parental cell line and EphA2 knock out (KO) A431 cell line. PVDF membrane was probed with 1 µg/mL of Goat Anti-Human EphA2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3035) followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). A specific band was detected for EphA2 at approximately 110 kDa (as indicated), but not detectable in the knockout A431 cell line. GAPDH (Catalog # MAB5718) is shown as a loading control. This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

A431 human epithelial carcinoma cell line was stained with Goat Anti-Human EphA2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3035, 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.

EphA2 was detected in immersion fixed paraffin-embedded sections of human ovarian cancer tissue using Goat Anti-Human EphA2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3035) at 5 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Goat HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS008) and counterstained with hematoxylin (blue). Specific labeling was localized to the plasma membrane of cancer cells. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

• Reactivity: Hu
• Applications: WB, Flow, IHC, CyTOF-ready, KO
• Clonality: Polyclonal
• Host: Goat
• Conjugate: Unconjugated
• Concentration: LYOPH

EphA2 Antibody [Unconjugated] Summary

• Immunogen: Mouse myeloma cell line NS0-derived recombinant human EphA2
  Gln25-Asn534
  Accession # P29317
• Specificity: Detects human EphA2 in direct ELISAs and Western blots. In direct ELISAs, less than 45% cross-reactivity with recombinant mouse EphA2 is observed and less than 1% cross-reactivity with recombinant human (rh) EphA1, rhEphA3, rhEphA4, rhEphA5, rhEphA6, rhEphA7 and rhEphA10 is observed.
• Source: N/A
• Isotype: IgG
• Clonality: Polyclonal
• Host: Goat
• Gene: EPHA2
• Purity Statement: Antigen Affinity-purified

Applications/Dilutions

• Dilutions:
  • CyTOF-ready
  • Flow Cytometry 0.25 ug/10^6 cells
  • Immunohistochemistry 5-15 ug/mL
  • Knockout Validated 1 ug/mL
  • Western Blot 1 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 either lyophilized or 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 EphA2 Antibody [Unconjugated]

• ARCC2
• EC 2.7.10
• EC 2.7.10.1
• Eck
• ECKepithelial cell receptor protein tyrosine kinase
• EPH receptor A2
• EphA2
• ephrin type-A receptor 2
• Epithelial cell kinase
• Myk2
• Sek2
• soluble EPHA2 variant 1
• Tyrosine-protein kinase receptor ECK

Background

EphA2, also known as Eck, Myk2, and Sek2, is a member of the Eph receptor tyrosine kinase family which binds Ephrins A1, 2, 3, 4, and 5 (1-4). A and B class Eph proteins have a common structural organization. The human EphA2 cDNA encodes a 976 amino acid (aa) precursor including a 24 aa signal sequence, a 510 aa extracellular domain (ECD), a 24 aa transmembrane segment, and a 418 aa cytoplasmic domain. The ECD contains an N-terminal globular domain, a cysteine-rich domain, and two fibronectin type III domains (5). The cytoplasmic domain contains a juxtamembrane motif with two tyrosine residues, which are the major autophosphorylation sites, a kinase domain, and a sterile alpha motif (SAM) (5). The ECD of human EphA2 shares 90-94% aa sequence identity with mouse, bovine, and canine EphA2, and approximately 45% aa sequence identity with human EphA1, 3, 4, 5, 7, and 8. EphA2 becomes autophosphorylated following ligand binding (6, 7) and then interacts with SH2 domain-containing PI3-kinase to activate MAPK pathways (8, 9). Reverse signaling is also propagated through the Ephrin ligand. Transcription of EphA2 is dependent on the expression of E-Cadherin (10), and can be induced by p53 family transcription factors (11). EphA2 is upregulated in breast, prostate, and colon cancer vascular endothelium. Its ligand, EphrinA1, is expressed by the local tumor cells (12, 13). In some cases, EphA2 and EphrinA1 are expressed on the same blood vessels (14). EphA2 signaling cooperates with VEGF receptor signaling in promoting endothelial cell migration (13). The gene encoding human EphA2 maps to a region on chromosome 1 which is frequently deleted in neuroectodermal tumors (15).

1. Poliakov, A. et al. (2004) Dev. Cell 7:465.
2. Surawska, H. et al. (2004) Cytokine Growth Factor Rev. 15:419. 
3. Pasquale, E.B. (2005) Nat. Rev. Mol. Cell Biol. 6:462.
4. Davy, A. and P. Soriano (2005) Dev. Dyn. 232:1.
5. Bohme, B et al. (1993) Oncogene 8:2857.
6. Pandey, A. et al. (1995) Science 268:567.
7. Bartley, T.D. et al. (1994) Nature 368:558.
8. Pandey, A. et al. (1994) J. Biol. Chem. 269:30154.
9. Miao, H. et al. (2001) Nat. Cell Biol. 3:527.
10. Orsulic, S. and R. Kemler (2000) J. Cell Sci. 113:1793.
11. Dohn, M. et al. (2001) Oncogene 20:6503.
12. Zelinski, D.P. et al. (2001) Cancer Res. 61:2301.
13. Brantley, D.M. et al. (2002) Oncogene 21:7011.
14. Ogawa, K. et al. (2000) Oncogene 19:6043.
15. Sulman, E.P. et al. (1997) Genomics 40:371.

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 EphA2 Antibody (AF3035)(10)

Publications using AF3035

• J Jukonen, L Moyano-Gal, K Höpfner, EA Pietilä, L Lehtinen, K Huhtinen, E Gucciardo, J Hynninen, S Hietanen, S Grénman, PM Ojala, O Carpén, K Lehti Aggressive and recurrent ovarian cancers upregulate ephrinA5, a non-canonical effector of EphA2 signaling duality Scientific Reports, 2021;11(1):8856. 2021 [PMID: 33893375] (Western Blot, Human)
• B Prinzing, P Schreiner, M Bell, Y Fan, G Krenciute, S Gottschalk MyD88/CD40 signaling retains CAR T cells in a less differentiated state JCI Insight, 2020;5(21):. 2020 [PMID: 33148882] (Western Blot, Human)
• L Moyano-Gal, EA Pietilä, SP Turunen, S Corvigno, E Hjerpe, D Bulanova, U Joneborg, T Alkasalias, Y Miki, M Yashiro, A Chernenko, J Jukonen, M Singh, H Dahlstrand, JW Carlson, K Lehti Adaptive RSK-EphA2-GPRC5A signaling switch triggers chemotherapy resistance in ovarian cancer EMBO Mol Med, 2020;0(0):e11177. 2020 [PMID: 32115889] (ICC, Human)
• A Visioli, F Giani, N Trivieri, R Pracella, E Miccinilli, MG Cariglia, O Palumbo, A Arleo, F Dezi, M Copetti, L Cajola, S Restelli, V Papa, A Sciuto, TP Latiano, M Carella, D Amadori, G Gallerani, R Ricci, S Alfieri, G Pesole, AL Vescovi, E Binda Stemness underpinning all steps of human colorectal cancer defines the core of effective therapeutic strategies EBioMedicine, 2019;0(0):. 2019 [PMID: 31056474] (ICC, Human)
• MR Xu, PF Wei, MZ Suo, Y Hu, W Ding, L Su, YD Zhu, WJ Song, GH Tang, M Zhang, P Li Brucine Suppresses Vasculogenic Mimicry in Human Triple-Negative Breast Cancer Cell Line MDA-MB-231 Biomed Res Int, 2019;2019(0):6543230. 2019 [PMID: 30723742] (Western Blot, Human)
• E Reátegui, KE van der Vo, CP Lai, M Zeinali, NA Atai, B Aldikacti, FP Floyd, A H Khankhel, V Thapar, FH Hochberg, LV Sequist, BV Nahed, B S Carter, M Toner, L Balaj, D T Ting, XO Breakefiel, SL Stott Engineered nanointerfaces for microfluidic isolation and molecular profiling of tumor-specific extracellular vesicles Nat Commun, 2018;9(1):175. 2018 [PMID: 29330365] (Functional Assay, Human)
• Barquilla A, Lamberto I, Heynen-Genel S, Brill L, Pasquale E Protein kinase A can block EphA2 receptor-mediated cell repulsion by increasing EphA2 S897 phosphorylation. Mol Biol Cell, 0;27(17):2757-70. 0 [PMID: 27385333] (Western Blot, Human)
• Sugiyama N, Gucciardo E, Tatti O, Varjosalo M, Hyytiainen M, Gstaiger M, Lehti K EphA2 cleavage by MT1-MMP triggers single cancer cell invasion via homotypic cell repulsion. J Cell Biol, 2013;201(3):467-84. 2013 [PMID: 23629968] (Western Blot, Human)
• Gordon K, Kochkodan J, Blatt H, Lin S, Kaplan N, Johnston A, Swindell W, Hoover P, Schlosser B, Elder J, Gudjonsson J, Getsios S Alteration of the EphA2/Ephrin-A signaling axis in psoriatic epidermis. J Invest Dermatol, 2013;133(3):712-22. 2013 [PMID: 23190894] (IHC-Fr, Human)
• Gopal U, Bohonowych JE, Lema-Tome C A novel extracellular Hsp90 mediated co-receptor function for LRP1 regulates EphA2 dependent glioblastoma cell invasion. PLoS ONE, 2011;6(3):e17649. 2011 [PMID: 21408136] (Flow Cytometry, Human)

Bioinformatics

• Gene Symbol: EPHA2
• Uniprot:
  • P29317()