Estrogen Receptor (ER) Antibody Staining Protocol for Immunohistochemistry


Description: Recognizes a protein of 67kDa, which is identified as estrogen receptor (ER) alpha. The ER gene consists of more than 140kb of genomic DNA divided into 8 exons, being translated into a protein with six functionally discrete domains, labeled A through F. This antibody strongly stains the nucleus of epithelial cells in breast carcinomas. The ER is an important regulator of growth and differentiation in the mammary gland. Presence of ER in breast tumors indicates an increased likelihood of response to anti-estrogen (e.g. tamoxifen) therapy.


Primary Antibody

Name: Estrogen Receptor (ER) Antibody

Clone: SP1, Rabbit anti-Human

Supplier: Lab Vision

Catalog Number: RM-9101-S

Dilution: 1:200 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

Reagent: Mayer's Hematoxylin
Staining Time: 30 seconds

Staining Pattern: Nuclear
Images: Search image

Additional Information:
Tissue Type: Breast carcinomas, MCF-7 cells.
Fixation: Formalin fixed paraffin sections
Positive Control: Breast carcinomas, MCF-7 cells.
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


Not tested on frozen sections



1. Katoh AK, Stemmler N, Specht S, D'Amico F (1997) Immunoperoxidase staining for estrogen and progesterone receptors in archival formalin fixed, paraffin embedded breast carcinomas after microwave antigen retrieval. Biotech Histochem. 72(6):291-8. PubMed Abstract