Investigating AMPK Signaling in DCIS Cells

MCF10.DCIS cells were plated at sub confluent density in 60 mm or 100 mm dishes for western blotting experiments. The day after plating the cells, the regular growth medium was removed and cells were treated with the indicated dose of fluvastatin, aspirin and metformin for 48 h in low-glucose growth medium. Total cellular protein was subjected to SDS-PAGE and transferred to Hybond ECL nitrocellulose membranes (Sigma Aldrich), which were probed with AMPK, pAMPK (Thr 172), HMGCR, or loading control vinculin antibodies. Proteins were detected using an Odyssey Classic infrared imaging system (Li-Cor Biosciences) as described previously [14 (link)].