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Fusion protein expressed in E. coli corresponding to amino acids 2577-3056.
Our Abpromise guarantee covers the use of ab78 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Flow Cyt||Use 1-2µg for 106 cells.|
|ICC/IF||Use at an assay dependent concentration.|
|IHC-P||Use a concentration of 1 µg/ml.|
|WB||1/2000. Detects a band of approximately 350 kDa (predicted molecular weight: 350 kDa).|
|IP||Use a concentration of 1 - 10 µg/ml.|
ab78 staining ATM in Human U2OS cells by Immunocytochemistry/ Immunofluorescence.
Cells were fixed with paraformaldehyde, permeabilized with 0.5% NP40/PBS and blocked with 3% BSA for 1 hour at 25°C. Samples were incubated with primary antibody (1/100 in 3% BSA/PBS) for 12 hours at 4°C. An AlexaFluor®488-conjugated goat anti-rabbit polyclonal IgG (1/200) (ab150077) was used as the secondary antibody.
Simona Cavalieri, University of Turin, ItalyArrows denote the location of the 45kDa beta actin protein, and the 350kDa ATM protein.
Detection of human ATM protein using anti-ATM 2C1 monoclonal antibody (ab78) by western blot.
Detection of human ATM protein using anti-ATM 2C1 monoclonal antibody (ab78) by immunoprecipitation.
ab78 (2µg/ml) staining ATM in human colonic mucosa, using an automated system (DAKO Autostainer Plus). Using this protocol there is strong nuclear staining of mucosal epithelium and lymphocytes.
Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer EDTA pH 9.0 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako envision flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.
Overlay histogram showing HeLa cells stained with ab78 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab78, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HeLa cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.
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