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Our Abpromise guarantee covers the use of ab9474 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 at an assay dependent concentration.
ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
|IHC-P||1/25 - 1/50. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.|
Upper band at 100-110KD corresponds to the "wild type AR." Lower band 70-75 KDa corresponds to a "truncated AR".
Blocking agent: 3% BSA, Dilution Buffer: TBST+3%BSA.
Upper band at 100-110KD corresponds to the wild type Androgen Receptor. Lower band 70-75 KDa corresponds to a truncated Androgen Receptor.
ab9474 staining human prostate by IHC-P.
ab9474 staining the Androgen Receptor in the human 22Rv1 Prostate Cancer Cell Line by ICC/IF (immunocytochemistry/immunofluorescence). Cells were formaldehyde fixed, permeabilized by 0.1% Triton for 15 minutes, and blocked with glycine (7.5mg/ml) for 15 minutes at room temperature. The sample was incubated with the primary antibody (1/50 in PBS) for 12 hours at 4°C. An undiluted Alexa Fluor 488®-conjugated Donkey anti-mouse polyclonal (ab150105) was used as the secondary.
Overlay histogram showing PC3 cells stained with ab9474 (red line). The cells were fixed with 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 (ab9474, 1/10 dilution) for 30 min at 22°C. The secondary antibody used was DyLight ® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. 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 decreased signal in PC3 cells fixed with 4% paraformaldehyde (10 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.
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