MCF7 cells were incubated at 37&degC for 1h with vehicle control (0 &microM) and different concentrations of rapamycin (ab120224). Decreased expression of mTOR (phospho S2448) (ab51044) in MCF7 cells correlates with an increase in rapamycin concentration, as described in literature.Whole cell lysates were prepared with RIPA buffer (containing protease inhibitors and sodium orthovanadate), 30&microg 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 ab51044 at 1 &microg/ml and ab8227 at 1 &microg/ml overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP (ab97051) at 1/10000 dilution and visualised using ECL development solution.

MCF7 cells were incubated at 37°C for 1h with vehicle control (0 µM) and different concentrations of rapamycin (ab120224). Decreased expression of mTOR (phospho S2448) (ab51044) in MCF7 cells correlates with an increase in rapamycin concentration, as described in literature.Whole cell lysates were prepared with RIPA buffer (containing protease inhibitors and sodium orthovanadate), 30µg 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 ab51044 at 1 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 dilution and visualised using ECL development solution.


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