Vimentin Antibody Staining Protocol for Immunohistochemistry


Description: Vimentin is a 55 kDa intermediate filament (IF) protein, which form part of the cytoskeleton of vertebrate cells. Among the five classes of IFs vimentin belongs to class III and is expressed in the cytoplasma of cells of mesenchymal origin. Being the predominant IF protein in mesenchymal cells vimentin was initially believed to be a useful marker for distinguishing non-epithelial from epithelial derived nepolasms. The co-expression of intermediate filaments, particularly vimentin and cytokeratin, has now been demonstrated in a variety of normal cells/tissues and in neoplastic lesions, necessitating the use of a panel of antibodies in differential tumour diagnosis.


Primary Antibody

Name: Mouse Anti-Human Vimentin Antibody

Clone: Vim 3B4

Supplier: DakoCytomation

Catalog Number: M7020

Dilution: 1:800 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
Incubation Time/Temperature: 1-3 minutes/room temperature

Reagent: Mayer's Hematoxylin
Staining Time: 30 seconds

Staining Pattern: Cytoplasmic
Images: Search image

Additional Information:
Species Reactivity: Human
Fixation: Formalin fixed paraffin sections
Positive Control: Normal breast
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. if IHC-TekTM Antibody Diluent (Cat# IW-1000 or IW-1001) is used, there is no serum blocking step needed.