BAG-1 (RAP46 Protein) Antibody Staining Protocol for Immunohistochemistry


Description: BAG-1, also called RAP46, is a multifunctional protein that regulates several cellular processes including cell survival, signal transduction and protein folding. BAG-1 is an anti-apoptotic protein capable of interacting with several types of proteins including those of the bcl-2 family, Raf-1, certain tyrosine kinase growth factor receptors and steroid hormone receptors. Alternative translation initiation points in the BAG-1 transcript results in the expression of four protein isoforms known as p50, p46, p33 and p29. The protein is expressed in a number of cell lines and has been demonstrated immunohistochemically to stain both the cytoplasm and nuclei of normal breast tissue.


Primary Antibody

Name: BAG-1 (RAP46 Protein) Antibody

Clone: 5C5, Mouse Anti-Human

Supplier: Novocastra Labs

Catalog Number: NCL-BAG-1

Dilution: 1:25 - 1:50 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: 60min/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: 95-100 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

Reagent: Mayer's Hematoxylin
Staining Time: 30 seconds

Staining Pattern: Nuclear and cytoplasmic
Images: Search image

Additional Information:
Tissue Type: Tonsil
Fixation: Formalin-fixed paraffin sections
Positive Control: Tonsil
Negative Control: Omit primary antibody, isotype control, 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