Ki67 Antibody Staining Protocol for Immunohistochemistry


Description: Ki-67 is a nuclear protein, which is expressed in proliferating cells. Ki-67 is preferentially expressed during late G1-, S-, M-, and G2-phases of the cell cycle, while cells in the G0 (quiescent) phase are negative for this protein.


Primary Antibody

Name: Ki67 Antibody

Clone: SP6, Rabbit monoclonal

Supplier: Lab Vision

Catalog Number: RM-9106-S1

Dilution: 1:200 - 1:500 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: Nuclear
Images: Search image

Additional Information:
Species Reactivity: Human, rat, mouse, dog and swine. Others not tested.
Fixation: Formalin fixed paraffin sections
Positive Control: Spleen, Tonsil, breast carcinomas, GI tract.
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