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CD45 Antibody Staining Protocol for Immunohistochemistry
Description: This antibody reacts with both alloantigens and all isoforms of the CD45 leukocyte common antigen (LCA), also known as Ly-5 or T200, found on all cells of hematopoietic origin, except erythrocytes. CD45 is a transmembrane glycoprotein which is expressed at high levels on the cell surface, and its presence distinguishes leukocytes from non-hematopoietic cells. The CD45 isoforms play complex roles in T-cell and B-cell antigen receptor signal transduction. This antibody is specific for mouse CD45 and suitable for immunohistochemical staining of Zinc-fixed and formalin fixed paraffin sections as well as acetone-fixed, frozen sections.
Primary Antibody
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Name: CD45 Antibody |
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Clone: 30-F11, Rat anti-Mouse |
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Supplier: BD Pharmingen |
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Catalog Number: 550539 |
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Dilution: 1:20 using IHC-TekTM Antibody Diluent (Cat# IW-1000 or IW-1001) to reduce background and unspecific staining and serum blocking step is NOT needed. |
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Incubation Time/Temp: 60min/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 |
Counterstain
| Reagent: Mayer's Hematoxylin |
| Staining Time: 30 seconds |
Results:
| Staining Pattern: Membrane |
| Images: Search image |
Additional Information:
| Species Reactivity: Mouse |
| Fixation: Formalin fixed paraffin sections, or acetone fixed frozen sections |
| Positive Control: Thymus, spleen |
| 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 need to perform antigen retrieval on acetone fixed frozen sections |
References: