Myeloperoxidase (MPO) Antibody Staining Protocol for Immunohistochemistry


Description: Myeloperoxidase (MPO), molecular weight 118 kDa, is a major constituent of cytoplasmic primary (azurophilic) granules of neutrophilic myeloid cells. The functional role of myeloperoxidase and other enzymes contained in primary granules is destruction of microorganisms and debris within the phagocytic vacuole of the cell. Since primary granules appear early in myeloid cell differentiation and are present throughout maturation, myeloperoxidase is a useful marker for for detection of myeloid cells.


Primary Antibody

Name: Myeloperoxidase Antibody

Clone: Rabbit anti-Human polyclonal

Supplier: Dako

Catalog Number: A0398

Dilution: 1:1000 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 (myeloid cells of both neutrophilic and eosinophilic types)
Images: Search image

Additional Information:
Tissue Type: Spleen
Fixation: Formalin fixed paraffin sections, acetone fixed frozen sections, or cell smears (no antigen retrieval needed for frozen sections or cell smears)
Positive Control: Spleen
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