TGase1 (Transglutaminase 1) Antibody Staining Protocol for Immunohistochemistry


Description: Terminally differentiating mammalian epidermal cells acquire an insoluble, 10 to 20 nm thick protein deposit on the intracellular surface of the plasma membrane known as the cross-linked cell envelope (CE). The CE is a component of the epidermis that is generated through formation of disulfide bonds and gamma-glutamyl-lysine isodipeptide bonds, which are formed by the action of transglutaminases (TGases). TGases are intercellularly localizing, Ca2+-dependent enzymes which catalyze the formation of isopeptide bonds by transferring an amine onto glutaminyl residues, thereby cross-linking glutamine residues and Lysine residues in substrate proteins. TGases influence numerous biological processes including blood coagulation, epidermal differentiation, seminal fluid coagulation, fertilization, cell differentiation and apoptosis.


Primary Antibody

Name: TGase1 (N-20)

Clone: Polyclonal Goat ant-Human

Supplier: Santa Cruz Biotechnology

Catalog Number: sc-18127

Dilution: 1:50 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 ºC - 100 ºC/room temperature
Heat/Cool Time: 20 minutes/20 minuters

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: Intercellular
Images: Search image

Additional Information:
Species Reactivity: Human, mouse, rat
Fixation: Formalin fixed paraffin sections.
Positive Control: Skin (epidermis)
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