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Our Abpromise guarantee covers the use of ab49314 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|ICC/IF||Use a concentration of 5 µg/ml.|
|WB||Use a concentration of 2.5 µg/ml. Detects a band of approximately 118 kDa (predicted molecular weight: 118 kDa).Can be blocked with Human Gli1 peptide (ab111659) or Human Gli1 peptide (ab86150). 5% skim milk in PBS buffer. (An HRP conjugated anti-Rabbit IgG could be used as a secondary, a suggested dilution of 1/50000 to 1/100000). Good results were obtained when blocked with 5% non-fat dry milk in 0.05% PBS-T.|
SKNSH cells were incubated at 37°C for 30 minutes with vehicle control (0 μM) and different concentrations of VU0155041 sodium salt (ab120248). Increased expression of Gli1 (ab49314) in SKNSH cells correlates with an decrease in VU0155041 sodium salt concentration, as described in literature.
Whole cell lysates were prepared with RIPA buffer (containing protease inhibitors and sodium orthovanadate), 30 μg of each were loaded on the gel and the WB was run under reducing conditions. After transfer the membrane was blocked for an hour using 5% BSA before being incubated with ab49314 at 2.5 μg/ml and ab8227 at 1 μg/ml overnight at 4°C. Antibody binding was detected using an anti- rabbit antibody conjugated to HRP (ab97051) at 1/10000 and visualised using ECL development solution.
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