Cytokeration 20 Antibody Staining Protocol for Immunohistochemistry


Description: Cytokeratins are intermediate filament cytoskeletal proteins essential to development and differentiation of epithelial cells. Approximately twenty different cytokeratins have been identified and are classified and numbered according to molecular weight and isoelectric points. In general, most low molecular weight cytokeratins (40-54 kDa) are distributed in nonsquamous epithelium, Moll’s catalog numbers 7-8 and/or 17-20. High molecular weight cytokeratins (48-67 kDa) are found in the squamous epithelium, Moll’s catalog numbers 1-6 and/or 9-16. CK20 (Clone KS20.8) is to identify epithelial cells in normal and neoplastic tissues using immunohistochemical methods. CK20 is valuable as a histodiagnostic tool for the subtyping of various carcinomas, including adenocarcinomas.1 It is an aid for a more precise classification of many epithelial tumors whose differential diagnosis is otherwise difficult. This antibody reacts with the 46 kDa cytokeratin intermediate filament protein as indicated by immunoblotting of cytokeratin-enriched cytoskeletal preparations isolated from villi of duodenal mucosa. The cytokeratin was initially described as “protein IT” but has recently been designated cytokeratin 201 supplementing the previously described number of the cytokeratin polypeptides. CK 20 is less acidic than the other type 1 cytokeratins and is of particular interest because of its restricted range of expression.


Primary Antibody

Name: Cytokeratin 20 Antibody

Clone: KS20.8, Mouse anti-Human

Supplier: Dako

Catalog Number: M7019

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: 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: 95-100 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:
Tissue Type: Skin
Fixation: Formalin fixed paraffin sections.
Positive Control: Intestinal epithelium, gastric foveolar epithelium, skin.
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.