EGFR Antibody Staining Protocol for Immunohistochemistry


Description: Epidermal growth factor receptor (EGFR) is a tyrosine protein kinase comprised of two different components of different molecular weights. Both components contain phosphotyrosine and phosphothreonine but only the higher molecular weight component contains phosphoserine. EGFR signaling involves small GTPases of the Rho family and EGFR trafficking involves small GTPases of the Rab family. EGFR expression is widespread in both normal and cancerous tissues.


Primary Antibody

Name: EGFR Antibody

Clone: Sheep Polyclonal

Supplier: LifeSpan BioSciences

Catalog Number: LS-B2914

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 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 Chromogen Substrate
Incubation Time/Temperature: 2-10 minutes/room temperature

Reagent: Mayer's Hematoxylin
Staining Time: 30 seconds

Staining Pattern: Cytoplasmic/membrane
Images: Search image

Additional Information:
Species Reactivity: Human, mouse, rat, chicken
Fixation: Formalin fixed paraffin sections
Positive Control: Human skin, placenta
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