Hypoxia-Inducible Factor 2 alpha (HIF-2-alpha) Antibody Staining Protocol for Immunohistochemistry

 

Description: Hypoxia contributes significantly to the pathophysiology of major categories of human disease, 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 humans and endothelial cells of the embryonic and adult mouse, whereas HIF-1alpha functions primarily in extravascular tissues. HIF-2 alpha is also a potent activator of the tie-2 gene, which is known to be selectively expressed in endothelial cells.

 

Primary Antibody

Name: HIF-2-alpha Antibody

Clone: Rabbit polyclonal

Supplier: Novus Biologicals

Catalog Number: NB100-122

Dilution: 1:300 - 1:500 using IHC-TekTM Antibody Diluent (Cat# IW-1000 or IW-1001) to reduce background and unspecific staining and serum blocking step is NOT needed.

Incubation Time/Temp: 60 minutes/room temperature


Antigen Retrieval
Device: IHC-TekTM Epitope Retrieval Steamer Set (Cat# IW-1102)
Buffer/pH value: IHC-TekTM Epitope Retrieval Solution (Cat# IW-1100)
Heat/Cool Temperature: 95 ºC - 100 ºC/room temperature
Heat/Cool Time: 20 minutes/20 minutes

Detection Methods
Standard Method: ABC Method or LSAB Method
Enhanced Method: Polymeric Methods

Chromogen Substrate
Reagent: DAB
Incubation Time/Temperature: 1-3 minutes/room temperature

Counterstain
Reagent: Mayer's Hematoxylin
Staining Time: 30 seconds

Results:
Staining Pattern: Nuclear/cytoplasmic
Images: Search image

Additional Information:

Species Reactivity: Mouse, rat, human

Fixation: Formalin fixed paraffin sections

Positive Control: Mouse skin

Negative Control: Omit primary antibody, isotype control or absorption control

Blocking: 2-5% normal serum to reduce unspecific background staining; 0.5-3% H2O2 to block endogenous peroxidase activity; avidin/biotin to block endogenous biotin activity if necessary


Notes:

Not tested on frozen sections.

 

References: