|Immunohistochemistry (Paraffin)||1:50 - 1:200|
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
NOTE: Do not allow slides to dry at any time during this procedure.
For Citrate: Heat slides in a microwave submersed in 1X citrate unmasking solution until boiling is initiated; follow with 10 min at a sub-boiling temperature (95°-98°C). Cool slides on bench top for 30 min.
|SignalStain® Boost IHC Detection Reagent (HRP, Rabbit) #8114||SignalStain® Boost IHC Detection Reagent (AP, Rabbit) #18653|
|SignalStain® DAB Substrate Kit #8059||SignalStain® Vibrant Red Alkaline Phosphatase Substrate Kit #76713|
|SignalStain® Vivid Purple Peroxidase Substrate Kit #96632||SignalStain® Ultra Blue Alkaline Phosphatase Substrate Kit #12824|
|SignalStain® Deep Black Peroxidase Substrate Kit #72986|
|SignalStain® Radiant Yellow Peroxidase Substrate Kit #69644|
NOTE: Use of detection reagents other than those specified in this protocol may require further optimization of the primary antibody to account for the different sensitivities of the detection reagents.
posted February 2010
revised June 2020
Protocol Id: 283
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Asp314 of mouse CD177 protein.
Cluster of differentiation 177 antigen (CD177, also known as human neutrophil alloantigen 2a and NB1), is a glycosyl-phosphatidylinositol (GPI)-linked N-glycosylated transmembrane protein expressed primarily on the surface of neutrophils (1,2). Expression of CD177 can be induced by granulocyte-colony-stimulating factor and during pregnancy or bacterial infection (3-5). CD177 mediates adhesion and transendothelial migration of neutrophils via interaction with CD31/PECAM-1 and integrin β2 on endothelial cells (6,7). CD177 also promotes the cell surface expression of proteinase 3 (PR3), which may play a role in enhancing endothelial cell junctional integrity and thus vascular integrity during neutrophil diapedesis (8-10). In association with CD11b/ITGAM, integrin β2, and PR3, CD177 promotes activation of TNF-α primed neutrophils including degranulation and superoxide production (11). CD177 is a polymorphic gene that has been linked to several important clinical diseases, including polycythemia vera, Wegener’s granulomatosis, and immune mediated neonatal neutropenia (12-14).
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