Human Tie-2 Antibody

Catalog # Availability Size / Price Qty
AF313
AF313-SP
Detection of Human Tie‑2 by Western Blot.
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Product Details
Citations (44)
FAQs
Supplemental Products
Reviews (1)

Human Tie-2 Antibody Summary

Species Reactivity
Human
Specificity
Detects human Tie-2 in direct ELISAs and Western blots.
Source
Polyclonal Goat IgG
Purification
Antigen Affinity-purified
Immunogen
Mouse myeloma cell line NS0-derived recombinant human Tie‑2
Ala23-Lys745
Accession # Q02763
Formulation
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.
Endotoxin Level
<0.10 EU per 1 μg of the antibody by the LAL method.
Label
Unconjugated

Applications

Recommended Concentration
Sample
Western Blot
1 µg/mL
See below
Simple Western
10 µg/mL
See below
Immunohistochemistry
5-15 µg/mL
See below
Blockade of Receptor-ligand Interaction
In a functional ELISA, 5-10 µg/mL of this antibody will block 50% of the binding of 40 ng/mL of biotinylated Recombinant Human Angiopoietin-2 (Catalog # BT623) to immobilized Recombinant Human Tie-2 Fc Chimera (Catalog # 313-TI) coated at 4 µg/mL (100 µL/well). At 50 μg/mL, this antibody will block >90% of the binding.
 

Please Note: Optimal dilutions should be determined by each laboratory for each application. General Protocols are available in the Technical Information section on our website.

Scientific Data

Western Blot Detection of Human Tie-2 antibody by Western Blot. View Larger

Detection of Human Tie‑2 by Western Blot. Western blot shows lysate of HUVEC human umbilical vein endothelial cells. PVDF membrane was probed with 1 µg/mL of Goat Anti-Human Tie-2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF313) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (HAF017). A specific band was detected for Tie-2 at approximately 150 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Immunohistochemistry Tie-2 antibody in Human Placenta by Immunohistochemistry (IHC-P). View Larger

Tie‑2 in Human Placenta. Tie-2 was detected in immersion fixed paraffin-embedded sections of human placenta using Goat Anti-Human Tie-2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF313) at 10 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Goat HRP-DAB Cell & Tissue Staining Kit (brown; CTS008) and counterstained with hematoxylin (blue). View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Simple Western Detection of Human Tie-2 antibody by Simple Western<SUP>TM</SUP>. View Larger

Detection of Human Tie‑2 by Simple WesternTM. Simple Western lane view shows lysates of HUVEC human umbilical vein endothelial cells, loaded at 0.2 mg/mL. A specific band was detected for Tie-2 at approximately 161 kDa (as indicated) using 10 µg/mL of Goat Anti-Human Tie-2 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF313) followed by 1:50 dilution of HRP-conjugated Anti-Goat IgG Secondary Antibody (HAF109). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.

Reconstitution Calculator

Reconstitution Calculator

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Preparation and Storage

Reconstitution
Reconstitute at 0.2 mg/mL in sterile PBS.
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Shipping
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. *Small pack size (SP) is shipped with polar packs. Upon receipt, store it immediately at -20 to -70 °C
Stability & 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.

Background: Tie-2

Tie-1/Tie (tyrosine kinase with Ig and EGF homology domains 1) and Tie-2/Tek comprise a receptor tyrosine kinase (RTK) subfamily with unique structural characteristics: two immunoglobulin-like domains flanking three epidermal growth factor (EGF)-like domains and followed by three fibronectin type III-like repeats in the extracellular region and a split tyrosine kinase domain in the cytoplasmic region. These receptors are expressed primarily on endothelial and hematopoietic progenitor cells and play critical roles in angiogenesis, vasculogenesis and hematopoiesis.

Human Tie-2 cDNA encodes a 1124 amino acid (aa) residue precursor protein with an 18 residue putative signal peptide, a 727 residue extracellular domain and a 354 residue cytoplasmic domain. Two ligands, angiopoietin-1 (Ang-1) and angiopoietin-2 (Ang-2), which bind Tie-2 with high-affinity have been identified. Ang-2 has been reported to act as an antagonist for Ang-1. Mice engineered to overexpress Ang-2 or to lack Ang-1 or Tie-2 display similar angiogenesis defects.

References
  1. Partanen, J. and D.J. Dumont (1999) Curr. Top. Microbiol. Immunol. 237:159.
  2. Takakura, N. et al. (1998) Immunity 9:677.
  3. Procopio, W. et al. (1999) J. Biol. Chem. 274:30196.
Long Name
Tyrosine Kinase with Immunoglobulin and Epidermal Growth Factor Homology Domains 2
Entrez Gene IDs
7010 (Human); 21687 (Mouse); 89804 (Rat); 396729 (Porcine); 403714 (Canine); 102122204 (Cynomolgus Monkey); 30747 (Zebrafish)
Alternate Names
angiopoietin-1 receptor; CD202b antigen; CD202b; EC 2.7.10; EC 2.7.10.1; hTIE2; p140 TEK; soluble TIE2 variant 1; soluble TIE2 variant 2; TEK tyrosine kinase, endothelial; TEK; Tie2; Tie-2; TIE2CD202b; Tunica interna endothelial cell kinase; Tyrosine-protein kinase receptor TEK; Tyrosine-protein kinase receptor TIE-2; venous malformations, multiple cutaneous and mucosal; VMCM; VMCM1

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Citations for Human Tie-2 Antibody

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

44 Citations: Showing 1 - 10
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  1. Circulating Tie2-Expressing Monocytes: A Potential Biomarker for Cervical Cancer
    Authors: Q Han, Q Zhang, F Ying, Z Wang, Y Zhang, L Gong, E Cai, J Qian, J Cai
    OncoTargets and Therapy, 2020-09-07;13(0):8877-8885.
  2. SMAD4 maintains the fluid shear stress set point to protect against arterial-venous malformations
    Authors: Kuheli Banerjee, Yanzhu Lin, Johannes Gahn, Julio Cordero, Purnima Gupta, Islam Mohamed et al.
    Journal of Clinical Investigation
  3. Polydom/SVEP1 binds to Tie1 and promotes migration of lymphatic endothelial cells
    Authors: Ryoko Sato-Nishiuchi, Masamichi Doiguchi, Nanami Morooka, Kiyotoshi Sekiguchi
    Journal of Cell Biology
  4. Lymphoid Lineage gamma δ T Cells Were Successfully Generated from Human Pluripotent Stem Cells via Hemogenic Endothelium
    Authors: Soo-Been Jeon, A-Reum Han, Yoo Bin Choi, Ah Reum Lee, Ji Yoon Lee
    International Journal of Stem Cells
  5. CD34dim cells identified as pluripotent stem cell‐derived definitive hemogenic endothelium purified using bone morphogenetic protein 4
    Authors: Soo‐Been Jeon, A‐Reum Han, Sunghun Lee, Seung Chan Lee, Min Ji Lee, Soon‐Jung Park et al.
    Cell Proliferation
  6. Increased frequency of proangiogenic tunica intima endothelial kinase 2 (Tie2) expressing monocytes in individuals with type 2 diabetes mellitus
    Authors: M Reijrink, J van Ark, CPH Lexis, LM Visser, ME Lodewijk, ICC van der Ho, CJ Zeebregts, H van Goor, SCA de Jager, G Pasterkamp, BHR Wolffenbut, JL Hillebrand
    Cardiovascular Diabetology, 2022-05-12;21(1):72.
    Species: Human
    Sample Types: Whole Tissue
    Applications: IHC
  7. Aberrant stromal tissue factor localisation and mycolactone-driven vascular dysfunction, exacerbated by IL-1beta, are linked to fibrin formation in Buruli ulcer lesions
    Authors: LT Hsieh, SJ Dos Santos, BS Hall, J Ogbechi, AD Loglo, FJ Salguero, MT Ruf, G Pluschke, RE Simmonds
    PloS Pathogens, 2022-01-31;18(1):e1010280.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot
  8. TNF-&alpha induces endothelial-mesenchymal transition promoting stromal development of pancreatic adenocarcinoma
    Authors: M Adjuto-Sac, P Soubeyran, J Garcia, S Audebert, L Camoin, M Rubis, J Roques, B Binétruy, JL Iovanna, R Tournaire
    Cell Death & Disease, 2021-06-25;12(7):649.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Immunoprecipitation
  9. Dynamic angiopoietin-2 assessment predicts survival and chronic course in hospitalized patients with COVID-19
    Authors: Erica Villa, Rosina Critelli, Simone Lasagni, Alessandra Melegari, Angela Curatolo, Ciro Celsa et al.
    Blood Advances
  10. Vanucizumab mode of action: Serial biomarkers in plasma, tumor, and skin-wound-healing biopsies
    Authors: F Heil, G Babitzki, A Julien-Laf, CH Ooi, M Hidalgo, C Massard, M Martinez-G, C Le Tournea, M Kockx, P Gerber, S Rossomanno, O Krieter, A Lahr, N Wild, SV Harring, K Lechner
    Translational Oncology, 2020-12-15;14(2):100984.
    Species: Human
    Sample Types: Whole Tissue
    Applications: IHC
  11. Sulfated glycans engage the Ang-Tie pathway to regulate vascular development
    Authors: ME Griffin, AW Sorum, GM Miller, WA Goddard, LC Hsieh-Wils
    Nat Chem Biol, 2020-10-05;0(0):.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot
  12. Role of the Ang2-Tie2 Axis in Vascular Damage Driven by High Glucose or Nucleoside Diphosphate Kinase B Deficiency
    Authors: A Chatterjee, R Eshwaran, H Huang, D Zhao, M Schmidt, T Wieland, Y Feng
    Int J Mol Sci, 2020-05-25;21(10):.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Immunoprecipitation
  13. Targeting the ER&beta/Angiopoietin-2/Tie-2 signaling-mediated angiogenesis with the FDA-approved anti-estrogen Faslodex to increase the Sunitinib sensitivity in RCC
    Authors: J Gu, Y Zhang, Z Han, L Gao, J Cui, Y Sun, Y Niu, B You, CP Huang, C Chang, X Wang, S Yeh
    Cell Death Dis, 2020-05-14;11(5):367.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot
  14. miR-15a/-16 Inhibit Angiogenesis by Targeting the Tie2 Coding Sequence: Therapeutic Potential of a miR-15a/16 Decoy System in Limb Ischemia
    Authors: Marie Besnier, Saran Shantikumar, Maryam Anwar, Parul Dixit, Aranzazu Chamorro-Jorganes, Walid Sweaad et al.
    Molecular Therapy - Nucleic Acids
  15. Weibel-Palade Bodies Orchestrate Pericytes During Angiogenesis
    Authors: Mélissande Cossutta, Marie Darche, Gilles Carpentier, Claire Houppe, Matteo Ponzo, Fabio Raineri et al.
    Arteriosclerosis, Thrombosis, and Vascular Biology
  16. Tie2 activation promotes choriocapillary regeneration for alleviating neovascular age-related macular degeneration
    Authors: J Kim, JR Park, J Choi, I Park, Y Hwang, H Bae, Y Kim, W Choi, JM Yang, S Han, TY Chung, P Kim, Y Kubota, HG Augustin, WY Oh, GY Koh
    Sci Adv, 2019-02-13;5(2):eaau6732.
    Species: Human, Mouse
    Sample Types: Cell Culture Supernates, Whole Cells, Whole Tissue
    Applications: ICC, IHC-Fr, Immunoprecipitation
  17. An acellular bioresorbable ultra-purified alginate gel promotes intervertebral disc repair: A preclinical proof-of-concept study
    Authors: T Tsujimoto, H Sudo, M Todoh, K Yamada, K Iwasaki, T Ohnishi, N Hirohama, T Nonoyama, D Ukeba, K Ura, YM Ito, N Iwasaki
    EBioMedicine, 2018-10-30;0(0):.
    Species: Rabbit
    Sample Types: Whole Cells
    Applications: ICC
  18. Structural basis of Tie2 activation and Tie2/Tie1 heterodimerization
    Authors: VM Leppänen, P Saharinen, K Alitalo
    Proc. Natl. Acad. Sci. U.S.A., 2017-04-10;0(0):.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot
  19. Endothelial destabilization by angiopoietin-2 via integrin beta1 activation.
    Authors: Hakanpaa L, Sipila T, Leppanen V, Gautam P, Nurmi H, Jacquemet G, Eklund L, Ivaska J, Alitalo K, Saharinen P
    Nat Commun, 2015-01-30;6(0):5962.
    Species: Human
    Sample Types: Whole Cells
    Applications: ICC, IHC
  20. Inhibition of Tyrosine Kinase Receptor Tie2 Reverts HCV-Induced Hepatic Stellate Cell Activation
    Authors: Samuel Martín-Vílchez, Yolanda Rodríguez-Muñoz, Rosario López-Rodríguez, Ángel Hernández-Bartolomé, María Jesús Borque-Iñurrita, Francisca Molina-Jiménez et al.
    PLoS ONE
  21. slanDCs selectively accumulate in carcinoma-draining lymph nodes and marginate metastatic cells.
    Authors: Vermi W, Micheletti A, Lonardi S, Costantini C, Calzetti F, Nascimbeni R, Bugatti M, Codazzi M, Pinter P, Schakel K, Tamassia N, Cassatella M
    Nat Commun, 2014-01-01;5(0):3029.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot
  22. Neurovascular Signals Suggest a Propagation Mechanism for Endogenous Stem Cell Activation Along Blood Vessels
    Authors: Jimmy Masjkur, Maria Adele Adele Rueger, Stefan R R. Bornstein, Ron McKay, Andreas Androutsellis-Theotokis
    CNS & Neurological Disorders - Drug Targets
  23. Chromatin structure of two genomic sites for targeted transgene integration in induced pluripotent stem cells and hematopoietic stem cells.
    Authors: van Rensburg, R, Beyer, I, Yao, X-Y, Wang, H, Denisenko, O, Li, Z-Y, Russell, D W, Miller, D G, Gregory, P, Holmes, M, Bomsztyk, K, Lieber, A
    Gene Ther, 2012-03-22;20(2):201-14.
    Species: Human
    Sample Types: Whole Cells
    Applications: ICC
  24. Exhaustion of nucleus pulposus progenitor cells with ageing and degeneration of the intervertebral disc.
    Authors: Sakai, Daisuke, Nakamura, Yoshihik, Nakai, Tomoko, Mishima, Taishi, Kato, Shunichi, Grad, Sibylle, Alini, Mauro, Risbud, Makarand, Chan, Danny, Cheah, Kathryn, Yamamura, Ken-ichi, Masuda, Koichi, Okano, Hideyuki, Ando, Kiyoshi, Mochida, Joji
    Nat Commun, 2012-01-01;3(0):1264.
    Species: Human
    Sample Types: Whole Cells
    Applications: Neutralization
  25. Opposing effects of the angiopoietins on the thrombin-induced permeability of human pulmonary microvascular endothelial cells.
    Authors: van der Heijden M, van Nieuw Amerongen GP, van Bezu J, Paul MA, Groeneveld AB, van Hinsbergh VW
    PLoS ONE, 2011-08-15;6(8):e23448.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Immunoprecipitation, Western Blot
  26. Tumour vascularization via endothelial differentiation of glioblastoma stem-like cells.
    Authors: Ricci-Vitiani L, Pallini R, Biffoni M, Todaro M, Invernici G, Cenci T, Maira G, Parati EA, Stassi G, Larocca LM, De Maria R
    Nature, 2010-11-21;468(7325):824-8.
    Species: Human
    Sample Types: Whole Cells, Whole Tissue
    Applications: Flow Cytometry, IHC-P
  27. Angiopoietin-2 interferes with anti-VEGFR2- induced vessel normalization and survival benefit in mice bearing gliomas
    Authors: Sung-Suk Chae, Walid S. Kamoun, Christian T. Farrar, Nathaniel D. Kirkpatrick, Elisabeth Niemeyer, Annemarie M.A. de Graaf et al.
    Clinical Cancer Research
  28. Cholera Toxin Regulates a Signaling Pathway Critical for the Expansion of Neural Stem Cell Cultures from the Fetal and Adult Rodent Brains
    Authors: Andreas Androutsellis-Theotokis, Stuart Walbridge, Deric M. Park, Russell R. Lonser, Ronald D. G. McKay
    PLoS ONE
  29. VEGF-DdeltaNdeltaC mediated angiogenesis in skeletal muscles of diabetic WHHL rabbits.
    Authors: Roy H, Bhardwaj S, Babu M, Lahteenvuo JE, Yla-Herttuala S
    Eur. J. Clin. Invest., 2010-05-01;40(5):422-32.
    Species: Rabbit
    Sample Types: Whole Tissue
    Applications: IHC-P
  30. The angiogenic factor angiopoietin-1 is a proneurogenic peptide on subventricular zone stem/progenitor cells.
    Authors: Rosa AI, Goncalves J, Cortes L, Bernardino L, Malva JO, Agasse F
    J. Neurosci., 2010-03-31;30(13):4573-84.
    Species: Mouse
    Sample Types: Cell Lysates, Whole Cells, Whole Tissue
    Applications: ICC, IHC, Neutralization, Western Blot
  31. Expression of VEGF receptors VEFGR-1 and VEGFR-2, angiopoietin receptors Tie-1 and Tie-2 in chorionic villi tree during early pregnancy
    Authors: Ramazan Demir
    Folia Histochemica et Cytobiologica
  32. Primary human acute myelogenous leukemia cells release matrix metalloproteases and their inhibitors: release profile and pharmacological modulation.
    Authors: Reikvam H, Hatfield KJ, Oyan AM, Kalland KH, Kittang AO, Bruserud O
    Eur. J. Haematol., 2009-11-17;84(3):239-51.
    Species: Human
    Sample Types: Whole Cells
    Applications: Neutralization
  33. Interaction with human stromal cells enhances CXCR4 expression and engraftment of cord blood Lin(-)CD34(-) cells.
    Authors: Kobune M, Kawano Y, Takahashi S, Takada K, Murase K, Iyama S, Sato T, Takimoto R, Niitsu Y, Kato J
    Exp. Hematol., 2008-06-17;36(9):1121-31.
    Species: Human
    Sample Types: Whole Cells
    Applications: Neutralization
  34. Angiopoietins assemble distinct Tie2 signalling complexes in endothelial cell-cell and cell-matrix contacts.
    Authors: Saharinen P, Eklund L, Miettinen J, Wirkkala R, Anisimov A, Winderlich M, Nottebaum A, Vestweber D, Deutsch U, Koh GY, Olsen BR, Alitalo K
    Nat. Cell Biol., 2008-04-20;10(5):527-37.
    Species: Human
    Sample Types: Whole Cells
    Applications: ICC, Western Blot
  35. Regulated proteolytic processing of Tie1 modulates ligand responsiveness of the receptor-tyrosine kinase Tie2.
    Authors: Marron MB, Singh H, Tahir TA, Kavumkal J, Kim HZ, Koh GY, Brindle NP
    J. Biol. Chem., 2007-08-29;282(42):30509-17.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot
  36. Interaction between Tie receptors modulates angiogenic activity of angiopoietin2 in endothelial progenitor cells.
    Authors: Kim KL, Shin IS, Kim JM, Choi JH, Byun J, Jeon ES, Suh W, Kim DK
    Cardiovasc. Res., 2006-08-07;72(3):394-402.
    Species: Human
    Sample Types: Cell Lysates, Whole Cells
    Applications: Immunoprecipitation, Neutralization, Western Blot
  37. Enhancement of angiogenic effectors through hypoxia-inducible factor in preterm primate lung in vivo.
    Authors: Asikainen TM, Waleh NS, Schneider BK, Clyman RI, White CW
    Am. J. Physiol. Lung Cell Mol. Physiol., 2006-05-05;291(4):L588-95.
    Species: Primate - Papio anubis (Olive Baboon)
    Sample Types: Tissue Homogenates
    Applications: Western Blot
  38. Spatial and temporal distribution of Tie-1 and Tie-2 during very early development of the human placenta.
    Authors: Kayisli UA, Cayli S, Seval Y, Tertemiz F, Huppertz B, Demir R
    Placenta, 2005-07-18;27(6):648-59.
    Species: Human
    Sample Types: Tissue Homogenates, Whole Tissue
    Applications: IHC-P, Western Blot
  39. Activation of hypoxia-inducible factors in hyperoxia through prolyl 4-hydroxylase blockade in cells and explants of primate lung.
    Authors: Asikainen TM, Schneider BK, Waleh NS, Clyman RI, Ho WB, Flippin LA, Gunzler V, White CW
    Proc. Natl. Acad. Sci. U.S.A., 2005-07-11;102(29):10212-7.
    Species: Human
    Sample Types: Whole Cells
    Applications: Western Blot
  40. Functional analysis of a mutant form of the receptor tyrosine kinase Tie2 causing venous malformations.
    Authors: Morris PN, Dunmore BJ, Tadros A, Marchuk DA, Darland DC, D&amp;apos;Amore PA, Brindle NP
    J. Mol. Med., 2004-10-29;83(1):58-63.
    Species: Human
    Sample Types: Cell Lysates, Whole Cells, Whole Tissue
    Applications: ICC, IHC, Western Blot
  41. Proangiogenic properties of human myeloma cells: production of angiopoietin-1 and its potential relationship to myeloma-induced angiogenesis.
    Authors: Giuliani N, Colla S, Lazzaretti M, Sala R, Roti G, Mancini C, Bonomini S, Lunghi P, Hojden M, Genestreti G, Svaldi M, Coser P, Fattori PP, Sammarelli G, Gazzola GC, Almici C, Caramatti C, Mangoni L, Rizzoli V
    Blood, 2003-03-20;102(2):638-45.
    Species: Human
    Sample Types: Whole Cells
    Applications: Neutralization
  42. Evidence for heterotypic interaction between the receptor tyrosine kinases TIE-1 and TIE-2.
    Authors: Marron MB, Hughes DP, Edge MD, Forder CL, Brindle NP
    J. Biol. Chem., 2000-12-15;275(50):39741-6.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Immunoprecipitation
  43. Structural insights into the clustering and activation of Tie2 receptor mediated by Tie2 agonistic antibody
    Authors: G Jo, J Bae, HJ Hong, AR Han, DK Kim, SP Hong, JA Kim, S Lee, GY Koh, HM Kim
    Nature Communications, 2021-11-01;12(1):6287.
  44. Presence of sst5TMD4, a truncated splice variant of the somatostatin receptor subtype 5, is associated to features of increased aggressiveness in pancreatic neuroendocrine tumors.
    Authors: Sampedro-Nunez M, Luque RM, Ramos-Levi AM et al.
    Oncotarget

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Human Tie-2 Antibody
By Anonymous on 10/17/2018
Application: ELISA Sample Tested: HASMC human aortic smooth muscle cells Species: Human

Binding activity of Tie2 antibody via antigen binding ELISA