 |
|
Factor VIII Antibody Staining Protocol for Immunohistochemistry
Description: Factor VIII is a glycoprotein present in human plasma. Factor VIII related antigen has been reported present in human endothelial cells. It is also present in megakaryocytes and platelets. Immunohistochemical staining for factor VIII related antigen could be used to determine if the benign and malignant neoplastic lesions are of endothelial origin. Since factor VIII related antigen is present in normal plasma and platelets, positive staining of intravascular plasma inevitably occurs in areas of hemorrhage or areas where vascular damage and leakage of plasma are present. Therefore, specific staining must be differentiated from non-specific background.
Primary Antibody
|
Name: Factor VIII Related Antigen Antibody |
|
Clone: Rabbit polyclonal |
|
Supplier: Zymed Labs |
|
Catalog Number: 18-0018 |
|
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: Incubator |
| Buffer/pH value: IHC-TekTM Proteinase K Solution (Cat# IW-1101) |
| Heat/Cool Temperature: 37 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 |
Counterstain
| Reagent: Mayer's Hematoxylin |
| Staining Time: 30 seconds |
Results:
| Staining Pattern: Membrane/cytoplasmic (endothelial cell) |
| Images: Search image |
Additional Information:
| Tissue Type: Tonsil, liver, kidney. |
| Fixation: Formalin fixed paraffin sections |
| Positive Control: Tonsil, liver, kidney. |
| 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 |
Notes:
Not tested on frozen sections |
References: