Hypoxia Induced Factor 1alpha

Mammalian cells require a constant supply of oxygen for survival. Low oxygen (hypoxia) leads to reduced oxidative phosphorylation and the depletion of cellular ATP that can result in cell death.  Hypoxia is a pleiotrophic condition generally encountered during embryonic development, organogenesis and in many pathological situations, like wound healing, inflammation, and tumorigenesis. Hypoxia induced apoptosis but also triggers adaptive mechanisms to ensure cell survival.  During tumor growth, the proliferating cells outstrip the vascular supply results in hypoxic environment that induces adaptive changes including induction of HIF1a and angiogenesis (1). Under hypoxic conditions activates the transcription of over 40 genes, including, erythropoietin, glucose transporters, glycolytic enzymes, vascular endothelial growth factor, and other genes whose protein products increase oxygen delivery or facilitate metabolic adaptation to hypoxiaThe post survival effects of HIF1a are mediated by induction of several genes including Neuron-derived Orphan receptor 1 (NOR1), insulin like growth factor binding protein 1 (IGFBP1), their transcription is up-regulated by HIF1a (2).  HIF1a is also one of the target of the mTOR pathway involved in longevity of lifespan specially in dietary restriction (3).   

At least two variants of HIF1a are cloned (HIF1a1 and HIF1A2) with 826 and 735 amino acids respectively.  HIF1a interacts with HIF1beta/ARNT subunit for DNA binding.  The interaction also promotes polyubiquitination and subsequent proteosomal degradation. During hypoxia, sumoylated HIF1A also binds VHL; the interaction promotes the ubiquitination of HIF1A. Interacts with SENP1; the interaction desumoylates HIF1A resulting in stabilization and activation of transcription.  HIF1a is expressed in most tissues with highest level in kidney and heart. HIF1a is  induced by many factors including growth factors. 

HIF1a is approximately 78 kDa protein. The HIF1a- antibodies were generated using peptide corresponding to human HIF1a1 and HIF1a1/2 proteins. HIF1a-selective antibodies are affinity purified over immobilized antigen based affinity chromatography, and the purified immunoglobulins are stabilized in antibody stabilization buffer. FabGennix Int. Inc. will provide limited quantities of antigenic blocking peptide/protein for competition assays involving HIF1a antibodies. Antibodies to HIF1a (HIF1-101AP) will label ~78kDa protein in Western blot positive control for HIF1a (PC-HIF1a) and several other cell lines.  FabGennix Inc. will conjugate this and other antibodies from its catalog to either secondary enzymes (alk-Pase or HRP) or fluorescent probes at a nominal cost upon request.  FabGennix also provides custom antibody production services for researchers that are looking for high affinity mono and polyclonal antibodies in various species. We specialize in making application specific antibodies that are useful in IHC, confocal and other applications where native antigen is detected. For a complete listing of all FabGennix antibodies please visit www.Fabgennix.com. 

Immunogen:          Synthetic peptides corresponding to unique epitope on HIF1A proteins, for HIF1-101AP epitope resides between aa 790-826; HIF1-112AP, between aa 150-200.  The peptide sequence selected was post synthetically modified to achieve desired antigenicity before covalently coupling to a carrier protein.

Concentration:      HIF1-101AP and HIF-112AP:  IgG concentration 0.55-0.57 mg/ml in antibody stabilization buffer.

Applications:         Antibodies HIF1-101AP and HIF-112AP are ideal for WB and immunoprecipitation, other applications have not been tested.  The species cross reactivity for these antibodies have not been examined fully.  WB: > 1:500; IMM & i.p pull-down assays: 1:200; IHC 1:200.  ELISA <1:10,000.  Application of this antibody in protocols not listed here does not necessarily exclude its use in such procedures. Investigators planning to use this antibody for applications not listed here can request a complimentary sample.

Reactivity:              This antibody detects a band of approximately ~78kDa in PC-HIF1a samples. There are some non-specific bands at 20-25kDa range, identity of these bands has not determined, but could represent proteolysis products.

Protocols:               Standard protocol for various applications (WB; IMM and IHC) of this antibody is provided with the product specification sheet; however, FabGennix Inc. strongly recommends investigators to optimize conditions for use of this antibody in their laboratories.  

Form/Storage:        The affinity-purified antibodies are isolated on immobilized antigen-affinity column and supplied as stabilized product.  Store at -20^oC for long-term storage.  FabGennix Inc. does not recommend storage of very dilute antibody solutions unless they are prepared in specially formulated multi use antibody dilution buffer (Cat # DiluOBuffer).  Working solutions of antibodies in DiluOBuffer should be filtered through 0.45um filter after every use for long-term storage.

 

Notes:  Briefly centrifuge to collect liquid, heat or boil PC-HIF1 tube for 1-2 minutes to dissolve any precipitate before use.  This product is “ready-to-use” for electrophoresis. After thawing store at room temperature, Repeated freezing and thawing may result in appearance of higher MW immunoreactive bands.   

 

New Reagents:  Now you can recycle your western blots (nitrocellulose, supported membranes and PVFD membranes) by using our StripObuffer (Cat FGI-1989).  Each stripping is guaranteed to give better signal (up to 8 stripping).  No strong pungent smell of reducing agents or heating required.  Block in 5X diluObuffer and you are ready for blotting with a new antibody

 

References:

1.         Greijer AE, van der Groep P, Kemming D, Shvarts A, Semenza GL, Meijer GA, van de Wiel MA, Belien JA, van Diest PJ, van der Wall E.  Up-regulation of gene expression by hypoxia is mediated predominantly by hypoxia-inducible factor 1 (HIF-1).  J Pathol. 2005 Jul;206(3):291-304.

2.         Lluis Martorell,† Maurizio Gentile,† Jordi Rius,‡ Cristina Rodríguez, Javier Crespo, Lina Badimon, and José Martínez-González. The Hypoxia-Inducible Factor 1/NOR-1 Axis Regulates the Survival Response of Endothelial Cells to Hypoxia.   Mol Cell Biol. 2009 November; 29(21): 5828–5842

3.         Di Chen, Emma Lynn Thomas, and Pankaj Kapahi.  HIF-1 Modulates Dietary Restriction-Mediated Lifespan Extension via IRE-1 in Caenorhabditis elegans.  PLoS Genet. 2009 May; 5(5): e1000486.

 

 

 

* For users who may require large amounts of Sec24D-401AP and FITC-Sec24D, please enquire about bulk material discounts.
This Product is for Research Use Only and is NOT intended for use in humans or clinical diagnosis.                                                                                                                                                                    031811-0020SF100 ge02-rev10.00

 

For research use only, not for diagnostic or therapeutic use.