The hypoxia-inducible transcription factor 2 alpha is stabilized in hypoxic tissue and, similarly to HIF-1 alpha, complexes with aryl hydrocarbon receptor nuclear translocator (ARNT). Both the HIF-1 and HIF-2 complexes bind hypoxia-response elements (HREs) in the promoters of many genes involved in adapting to an environment of insufficient oxygen or hypoxia. HIF-1 and HIF-2 do not appear to be completely redundant, although specific functions are only beginning to be elucidated. Hypoxic tissue environments occur in vascular and pulmonary diseases as well as cancer, indicating the potentially broad impact of gene regulation by both HIF-1 alpha and HIF-2 alpha.
Human/Rat Total HIF-2 alpha/EPAS1 DuoSet IC ELISA
R&D Systems | Catalog # DYC2997-2
Discontinued Product
DYC2997-2 has been discontinued.
View all HIF-2 alpha/EPAS1 products.
Key Product Details
Assay Type
Solid Phase Sandwich ELISA
Sample Type
Cell Lysates
Reactivity
Human, Rat
Human/Rat Total HIF-2 alpha/EPAS1 DuoSet IC ELISA Features
- Optimized capture and detection antibody pairings with recommended concentrations save lengthy development time
- Development protocols are provided to guide further assay optimization
- Assay can be customized to your specific needs
- Available in 2, 5, and 15- (96-well) plate pack sizes
- Economical alternative to Western blot
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Product Summary for Human/Rat Total HIF-2 alpha/EPAS1 DuoSet IC ELISA
This DuoSet IC ELISA contains the basic components required for the development of sandwich ELISAs to measure HIF-2 alpha / EPAS1 in cell lysates. An immobilized capture antibody specific for HIF-2 alpha / EPAS1 binds both phosphorylated and unphosphorylated HIF-2 alpha / EPAS1. After washing away unbound material, a biotinylated detection antibody is used to detect both phosphorylated and unphosphorylated protein, utilizing a standard Streptavidin-HRP format.
Product Specifications
Assay Format
96-well strip plate (sold separately)
Sample Volume Required
Cell lysates (100 µL)
Detection Method
Colorimetric ELISA - 450nm (TMB)
Conjugate
Biotin
Specificity
Label
HRP
Scientific Data Images for Human/Rat Total HIF-2 alpha/EPAS1 DuoSet IC ELISA
Standard Curve
Total HIF-2 alpha Detection in HepG2 Cell Lysate.
Lysate prepared from HepG2 human hepatocellular carcinoma cells treated with 150 μM of CoCl2 for 18 hours were incubated in wells coated with Total HIF-2 alpha Capture Antibody. Unbound material was removed by washing and bound material was solubilized in SDS gel sample buffer. The same lysate and captured proteins were electrophoresed and transferred to a PVDF membrane. The captured protein was immunoblotted with Total HIF-2 alpha Detection Antibody and the lysate was immunoblotted with anti-HIF-2 alpha antibody. Only one band corresponding to HIF-2 alpha was detected in the captured material.Cross-reactivity experiments were performed with this DuoSet IC ELISA to further determine specificity. Recombinant mouse HIF-2 alpha was assayed at 80 ng/mL and read 16.4 ng/mL (20.5% cross-reactivity). Recombinant human (rh) HIF-1 alpha and rhARNT were assayed at 80 ng/mL and did not cross-react or interfere in the assay.
Total HIF-1 alpha Detection in 786-O, Caki-2, and MCF-7 Cell Lysate.
Lysate prepared from 786-O human renal adenocarcinoma cells, Caki-2 human kidney clear carcinoma cells, and MCF-7 human breast cancer cells untreated or treated with 150 μM of CoCl2 for 18 hours were analyzed with both this DuoSet IC ELISA and the Human/Mouse Total HIF-1 alpha DuoSet IC ELISA (R&D Systems, Catalog # DYC1935). The 786-O cells are positive for HIF-2 alpha only while Caki-2 and MCF-7 treated with CoCl2 are positive for HIF-1 alpha only. Only the HIF-2 alpha positive 786-O cells were detected with the Human/Rat Total HIF-2 alpha /EPAS1 DuoSet IC ELISA, indicating that this DuoSet IC ELISA is specific for HIF-2 alpha.
Total HIF-2 alpha/EPAS1 Detection in 786-O and HepG2 Cell Lysate.
Lysates prepared from 786-O human renal adenocarcinoma cell and HepG2 human hepatocellular carcinoma cells untreated or treated with 150 μM of CoCl2 for 18 hours were quantified with this DuoSet IC ELISA. The same lysates were also immunoblotted (inset) with anti-HIF-2 alpha antibody. The DuoSet IC ELISA results correlated well with the relative amounts of HIF-2 alpha detected by Western blot.
Total HIF-2 alpha/EPAS1 Detection in PC-12 Cell Lysate.
Lysates prepared from PC-12 rat adrenal pheochromocytoma cells untreated or treated with 150 μM of CoCl2 for 18 hours were quantified with this DuoSet IC ELISA. The same lysates were also immunoblotted (inset) with anti-HIF-2 alpha antibody (R&D Systems, Catalog # AF2886). The DuoSet IC ELISA results correlate well with the relative amounts of HIF-2 alpha detected by Western blot.Kit Contents for Human/Rat Total HIF-2 alpha/EPAS1 DuoSet IC ELISA
- Capture Antibody
- Conjugated Detection Antibody
- Calibrated Immunoassay Standard or Control
- Streptavidin-HRP
Other Reagents Required
PBS: (Catalog # DY006), or 137 mM NaCl, 2.7 mM KCl, 8.1 mM Na2HPO4, 1.5 mM KH2O4, pH 7.2 - 7.4, 0.2 µm filtered
Wash Buffer: (Catalog # WA126), or equivalent
Lysis Buffer*
IC Diluent*
Blocking Buffer*
Substrate Solution: 1:1 mixture of Color Reagent A (H2O2) and Color Reagent B (Tetramethylbenzidine) (Catalog # DY999)
Stop Solution: 2 N H2SO4 (Catalog # DY994)
Microplates: From Costar EIA Plate (Costar Catalog # 2592) or R&D Systems (Catalog # DY990), or equivalent
Plate Sealers: ELISA Plate Sealers (Catalog # DY992), or equivalent
*For the Lysis Buffer, IC Diluent, and Blocking BUffer recommended for a specific DuoSet ELISA Development Kit, please see the product
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Preparation and Storage
Shipping
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.
Background: HIF-2 alpha/EPAS1
Long Name
Hypoxia-inducible Transcription Factor 2 alpha
Alternate Names
EPAS1, HIF 2A, HIF2 alpha, HIF2A, HLF, MOP2
Gene Symbol
EPAS1
Additional HIF-2 alpha/EPAS1 Products
Product Documents for Human/Rat Total HIF-2 alpha/EPAS1 DuoSet IC ELISA
Certificate of Analysis
To download a Certificate of Analysis, please enter a lot or batch number in the search box below.
Note: Certificate of Analysis not available for kit components.
Product Specific Notices for Human/Rat Total HIF-2 alpha/EPAS1 DuoSet IC ELISA
For research use only
Citations for Human/Rat Total HIF-2 alpha/EPAS1 DuoSet IC ELISA
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Protocols
Find general support by application which include: protocols, troubleshooting, illustrated assays, videos and webinars.
- ELISA Sample Preparation & Collection Guide
- ELISA Troubleshooting Guide
- How to Run an R&D Systems DuoSet ELISA
- How to Run an R&D Systems Quantikine ELISA
- How to Run an R&D Systems Quantikine™ QuicKit™ ELISA
- Quantikine HS ELISA Kit Assay Principle, Alkaline Phosphatase
- Quantikine HS ELISA Kit Principle, Streptavidin-HRP Polymer
- Sandwich ELISA (Colorimetric) – Biotin/Streptavidin Detection Protocol
- Sandwich ELISA (Colorimetric) – Direct Detection Protocol
- Troubleshooting Guide: ELISA
- View all Protocols, Troubleshooting, Illustrated assays and Webinars
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Associated Pathways
HIF Repressor Pathways
