Endothelial PAS Domain Protein 1 (EPAS1) (AA 535-631) antibody

Details for Product No. ABIN151063
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Antigen
Synonyms EPAS1, ECYT4, HIF2A, HLF, MOP2, PASD2, bHLHe73, HIF-2alpha, HRF, Hif2a, bhlhe73, ecyt4, hif2, hif2a, hif2alpha, hlf, mop2, pasd2, EPAS, EPAS-1
Epitope
AA 535-631
(32), (15), (7), (6), (6), (5), (5), (3), (2), (2), (2), (2), (2), (2), (2), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1)
Reactivity
Human
(106), (42), (30), (13), (13), (7), (1), (1), (1), (1)
Host
Mouse
(81), (47), (2), (1)
Clonality (Clone)
Monoclonal ()
Conjugate
Un-conjugated
(10), (5), (4), (4), (4), (4), (2), (2), (2), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1)
Application
Western Blotting (WB), Chromatin Immunoprecipitation (ChIP), ELISA, Flow Cytometry (FACS), Immunocytochemistry (ICC), Immunofluorescence (IF), Immunohistochemistry (IHC), Immunohistochemistry (Paraffin-embedded Sections) (IHC (p)), Immunoprecipitation (IP)
(114), (52), (43), (36), (36), (27), (16), (14), (10), (6), (3), (2), (1), (1)
Pubmed 15 references available
Quantity 0.1 mL
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Catalog No. ABIN151063
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Immunogen Human HIF-2 alpha, corresponding to amino acids 535-631.
Clone Ep190b
Isotype IgG1
Specificity This is specific for HIF-2 alpha and does not cross-react with HIF-1 alpha.
Cross-Reactivity (Details) Ability to use in mouse is mixed with some positive,some negative results.
Purification Protein G purified
Alternative Name HIF2A / HIF2 alpha
Background Hypoxia contributes significantly to the pathophysiology of major categories of humandisease, including myocardial and cerebral ischemia, cancer, pulmonary hypertension,congenital heart disease and chronic obstructive pulmonary disease.HIF-2 alpha is predominantly expressed in highly vascularized tissues of adult humansand endothelial cells of the embryonic and adult mouse, whereas HIF-1 alpha functionsprimarily in extravascular tissues. HIF-2 alpha is also a potent activator of the tie-2gene, which is known to be selectively expressed in endothelial cells. Alternate Names: anti-Endothelial pas domain protein 1 antibody, anti-Hif2a antibody, anti-Hypoxiainducible factor 2 alpha subunit antibody, anti-Member of pas superfamily 2 antibody,anti-MOP2 antibody, anti-EPAS1 antibody, anti-Hif-2a antibody, anti-Hif 2a antibody,anti-Hif 2 alpha antibody.
Gene Symbol: EPAS1
Gene ID 2034, 13819, 29452
UniProt Q99814
Application Notes This HIF-2 alpha (ep190b) antibody is useful for ELISA, Flow Cytometry, Immunohistochemistry on paraffin-embedded sections and Western Blot. In WB, it recognizes a band at ~118 kDa representing HIF-2 alpha.
Recommended dilutions: Chromatin Immunoprecipitation, ELISA 1:100-1:2000, Flow Cytometry 1:400, Immunocytochemistry/Immunofluorescence, Immunohistochemistry 1:150-1:300, Immunohistochemistry-Paraffin 1:150-1:300, Immunoprecipitation, Western Blot 1-2 µg/mL
Protocol Protocol specific for HIF-2 alpha Antibody Procedure Guide for NB 100-132Monoclonal Anti-HIF-2 alpha Western Blot Procedure
1. Resolve nuclear cell extracts (50-100 µg/lane) on a 6 % SDS-polyacrylamide gel, under reducing conditions.
. Transfer to a nitrocellulose membrane, overnight, or to a *PVDF membrane [*in 20 mM Tris/100 mM glycine/10 % (v/v) methanol/0.05 % SDS].
. Block the membrane in TBS containing 5 % non-fat dry milk and 0.1 % Tween-20.
. Rinse the membrane in TBST, twice.
. Incubate the membrane in anti-HIF-2 alpha , diluted 1:500 in TBS+1 % BSA, overnight at 4C.
. Wash membrane with TBST for 35 minutes at RT (1 X 15 minutes, 2 X 10 minutes).
. Incubate the membrane with diluted HRP conjugated goat anti-mouse antibody.
. Wash membrane with TBST for 35 minutes at RT (1 X 15 minutes, 2 X 10 minutes).
. Use Amersham ECL Kit, as directed, to detect image.Immunohistochemistry Procedure for Paraffin Sections
. Prior to performing the IPOX experiment, dewax the paraffin sections by baking them at 60C for 30 minutes and then putting them through citroclear.
. Hydrate the sections through the following series:A. 3 X 5 minutes xylenesB. 3 X 5 minutes 100 % EtohC. 2 minutes 95 % EtohD. 2 minutes 70 % EtohE. 1 minute 50 % EtohF. 1 minute ddH2OG. 1 minute TBS
. Block endogenous peroxidase with 0.5 % hydrogen peroxide in water, for 30 minutes.
. Antigen unmasking is performed by incubating at 60C for 16 hours, in 50mmol/L Tris and 0.2 mmol/L EDTA (pH 9.0), using a covered water bath.
. Rinse slides with PBS and then incubate with PBS containing 0.2 % Triton X-100 for 10 minutes.
. Rinse slides with PBS.
. Incubate sections with 1:1,000-1:3,000 dilution of anti-HIF-2 alpha for 90 minutes at RT.
. Incubate sections in secondary HRP-conjugated goat anti-mouse serum for 30 minutes at RT.
. Incubate sections in tertiary HRP-conjugated rabbit anti-goat serum for 30 minutes at RT.
. Develop the peroxidase reaction using diaminobenzidine.
. Wash slide and mount in aqueous mountant.Substitution of the primary antibody with PBS can be used as a negative control.
. Sub-confluent cells are grown on chamber slides and incubated for 16 hours either in air or under 0.1 % hypoxia.
. Wash cells in ice-cold PBS.
. Fix cells in formaldehyde (3.7 % in PBS) for 10 minutes at room temperature (RT).
. Wash cells twice, in PBS, and permeabilize by incubating in 0.2 % Triton X-100 in PBS for 10 minutes at RT.
. Incubate the slides with 1:1,000-1:3,000 dilution of anti-HIF-2 alpha for 1 hour at RT.
. Wash in PBS for 5 minutes.
. Incubate with HRP-conjugated goat anti-mouse for 30 minutes at RT.
. Detect binding using 3Prime-diaminobenzidine.
. Counterstain with hematoxylin.IHC-FFPE sectionsI. Deparaffinization: A. Treat slides with Xylene: 3 changes for 5 minutes each. Drain slides for 10 seconds between changes. B. Treat slides with 100 % Reagent Alcohol: 3 changes for 5 minutes each. Drain slides for 10 seconds between changes.II. Quench Endogenous Peroxidase: A. Place slides in peroxidase quenching solution: 15-30 minutes. To Prepare 200 mL of Quenching Solution: -Add 3 mL of 30 % Hydrogen Peroxide to 200 mL of Methanol.
Restrictions For Research Use only
Format Liquid
Concentration 1 mg/mL
Buffer PBS, Sodium Azide
Preservative Sodium azide
Precaution of Use WARNING: Reagents contain sodium azide. Sodium azide is very toxic if ingested or inhaled. Avoid contact with skin, eyes, or clothing. Wear eye or face protection when handling. If skin or eye contact occurs, wash with copious amounts of water. If ingested or inhaled, contact a physician immediately. Sodium azide yields toxic hydrazoic acid under acidic conditions. Dilute azide-containing compounds in running water before discarding to avoid accumulation of potentially explosive deposits in lead or copper plumbing.
Handling Advice Avoid freeze-thaw cycles
Storage -20 °C
Storage Comment Aliquot and store at -20 °C or -80 °C.
Supplier Images
anti-Endothelial PAS Domain Protein 1 (EPAS1) (AA 535-631) antibody HIF-2 alpha detected in hypoxic human lysate using ABIN151063. Lane 1: normoxic A549 lysate control, lane 2: hypoxic A549 lysate.
anti-Endothelial PAS Domain Protein 1 (EPAS1) (AA 535-631) antibody (2) Staining of heart, cardiac myocytes using ABIN151063.
General Wiesener, Turley, Allen et al.: "Induction of endothelial PAS domain protein-1 by hypoxia: characterization and comparison with hypoxia-inducible factor-1alpha." in: Blood, Vol. 92, Issue 7, pp. 2260-8, 1998 (PubMed).

Xia, Kageyama, Hayashi et al.: "Positive expression of HIF-2alpha/EPAS1 in invasive bladder cancer." in: Urology, Vol. 59, Issue 5, pp. 774-8, 2002 (PubMed).

Mohan, Burk: "von Hippel-Lindau protein complex is regulated by cell density." in: Oncogene, Vol. 22, Issue 34, pp. 5270-80, 2003 (PubMed).

Zimmer, Doucette, Siddiqui et al.: "Inhibition of hypoxia-inducible factor is sufficient for growth suppression of VHL-/- tumors." in: Molecular cancer research : MCR, Vol. 2, Issue 2, pp. 89-95, 2004 (PubMed).

Pan, Oprysko, Asham et al.: "p53 cannot be induced by hypoxia alone but responds to the hypoxic microenvironment." in: Oncogene, Vol. 23, Issue 29, pp. 4975-83, 2004 (PubMed).

Koike, Kimura, Miyazaki et al.: "Hypoxia induces adhesion molecules on cancer cells: A missing link between Warburg effect and induction of selectin-ligand carbohydrates." in: Proceedings of the National Academy of Sciences of the United States of America, Vol. 101, Issue 21, pp. 8132-7, 2004 (PubMed).

Harvey, Kind, Pantaleon et al.: "Oxygen-regulated gene expression in bovine blastocysts." in: Biology of reproduction, Vol. 71, Issue 4, pp. 1108-19, 2004 (PubMed).

Mekhail, Gunaratnam, Bonicalzi et al.: "HIF activation by pH-dependent nucleolar sequestration of VHL." in: Nature cell biology, Vol. 6, Issue 7, pp. 642-7, 2004 (PubMed).

Grabmaier, A de Weijert, Verhaegh et al.: "Strict regulation of CAIX(G250/MN) by HIF-1alpha in clear cell renal cell carcinoma." in: Oncogene, Vol. 23, Issue 33, pp. 5624-31, 2004 (PubMed).

Rathmell, Hickey, Bezman et al.: "In vitro and in vivo models analyzing von Hippel-Lindau disease-specific mutations." in: Cancer research, Vol. 64, Issue 23, pp. 8595-603, 2004 (PubMed).

Yoshimura, Dhar, Kohno et al.: "Prognostic impact of hypoxia-inducible factors 1alpha and 2alpha in colorectal cancer patients: correlation with tumor angiogenesis and cyclooxygenase-2 expression." in: Clinical cancer research : an official journal of the American Association for Cancer Research, Vol. 10, Issue 24, pp. 8554-60, 2004 (PubMed).

Nakamura, Abreu-e-Lima, Awakura et al.: "Clusterin is a secreted marker for a hypoxia-inducible factor-independent function of the von Hippel-Lindau tumor suppressor protein." in: The American journal of pathology, Vol. 168, Issue 2, pp. 574-84, 2006 (PubMed).

Zheng, Ruas, Cao et al.: "Cell-type-specific regulation of degradation of hypoxia-inducible factor 1 alpha: role of subcellular compartmentalization." in: Molecular and cellular biology, Vol. 26, Issue 12, pp. 4628-41, 2006 (PubMed).

Koizume, Jin, Miyagi et al.: "Activation of cancer cell migration and invasion by ectopic synthesis of coagulation factor VII." in: Cancer research, Vol. 66, Issue 19, pp. 9453-60, 2006 (PubMed).

Bernhardt, Wiesener, Weidemann et al.: "Involvement of hypoxia-inducible transcription factors in polycystic kidney disease." in: The American journal of pathology, Vol. 170, Issue 3, pp. 830-42, 2007 (PubMed).

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