Description: Apoptosis is one of a number of phenotypic responses that may occur as a result of signal transduction pathways occurring in the cell. One identified mechanism for initiating caspase activation requires the participation of mitochondria and involves apoptosis protease activating factor-1 (Apaf-1). Apaf-1 is a cytosolic protein that rests in a latent state until bound by cytochrome c (Apaf-2). Cytochrome c is commonly released from the mitochondria during apoptosis induced by many but probably not all cell death stimuli. The resulting Apaf1/cytochrome c complex associates with the zyymogen form of caspase-9 (Apaf-3) in the presence of dATP or ATP, promoting the autocatalytic activation of caspase-9. Once activated caspase-9 can then cleave and activate procaspase-3 directly, resulting in a cascade of additional caspase activation and apoptosis.
Primary Antibody
Name: Apoptosis Protease Activating Factor 1 Antibody |
Clone: Rabbit polyclonal |
Supplier: Novocastra Labs |
Catalog Number: NCL-APAF1 |
Dilution: 1:20 - 1:40 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: 60min/room temperature |
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 |
Standard Method: ABC Method or LSAB Method |
Enhanced Method: Polymeric Methods |
Reagent: DAB |
Incubation Time/Temperature: 1-3 minutes/room temperature |
Reagent: Mayer's Hematoxylin |
Staining Time: 30 seconds |
Staining Pattern: Cytoplasmic |
Images: Search image |
Tissue Type: Myocardium or skin, testis |
Fixation: Formalin-fixed paraffin sections, or acetone fixed frozen sections |
Positive Control: Myocardium or skin, testis |
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 |
There is no need to perform antigen retrieval on acetone fixed frozen sections |
References: