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Androgen Receptor Antibody Staining Protocol for Immunohistochemistry
Description: Androgen receptor is a member of the superfamily of ligand responsive transcription regulators. The androgen receptor functions in the nucleus where it is believed to act as a transcriptional regulator mediating the action of male sex hormones (androgens). The androgen receptor has wide tissue distribution and can be demonstrated by immunohistochemistry in several tissues eg prostate (Zhuang et al., 1992), skin (Blauer et al., 1991) and oral mucosa.
Primary Antibody
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Name: Androgen Receptor Antibody |
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Clone: 2F12, Mouse Anti-Human |
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Supplier: Novocastra Labs |
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Catalog Number: NCL-APP |
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Dilution: 1:25 using IHC-TekTM Antibody Diluent (Cat# IW-1000 or IW-1001) to reduce background and unspecific staining and serum blocking step is NOT needed. |
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Incubation Time/Temp: 60 min/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 |
Counterstain
| Reagent: Mayer's Hematoxylin |
| Staining Time: 30 seconds |
Results:
| Staining Pattern: Nuclear |
| Images: Search image |
Additional Information:
| Tissue Type: Prostate |
| Fixation: Formalin-fixed paraffin sections, or Zamboni’s-fixed, frozen sections. |
| Positive Control: Prostate |
| 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 |
Notes:
There is no need to perform antigen retrieval on frozen sections |
References:
1. Braidman IP, Baris C, Selby PL, Adams JE, Freemont AJ, Hoyland JA. (2000) Preliminary report of impaired oestrogen receptor-alpha expression in bone, but no involvement of androgen receptor, in male idiopathic osteoporosis. J Pathol. 192(1):90-6. PubMed Abstract
2. Stefanini M, De Martino C, Zamboni L. (1967) Fixation of ejaculated spermatozoa for electron microscopy. Nature. 1967 Oct 14;216(111):173-4. PubMed Abstract.
3. Magi-Galluzzi C, Montironi R, Prete E, Kwan PW, Delellis RA (1996) Effect of microwave oven heating times on androgen receptor antigen retrieval from paraffin-embedded prostatic adenocarcinoma. Anticancer Res. 16(5A):2931-6. PubMed Abstract