Bcl-2 Oncoprotein Antibody Staining Protocol for Immunohistochemistry


Description: Bcl-2 is a member of a family of proteins that are involved in apoptosis. Bcl-2 is an integral inner mitochondrial membrane protein of 25kD and has a wide tissue distribution. It is considered to act as an inhibitor of apoptosis. For this reason, bcl-2 expression is inhibited in germinal centres where apoptosis forms part of the B cell production pathway.


Primary Antibody

Name: Bcl-2 Oncoprotein Antibody

Clone: 3.1, Mouse Anti-Human

Supplier: Novocastra Labs

Catalog Number: NCL-bcl-2-486

Dilution: 1:100 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: Membrane 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



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2. Takes RP, Baatenburg de Jong RJ, Wijffels K, Schuuring E, Litvinov SV, Hermans J, van Krieken JH. (2001) Expression of genetic markers in lymph node metastases compared with their primary tumours in head and neck cancer. J Pathol. 194(3):298-302. PubMed Abstract.