DMEM–Dulbecco's Modified Eagle Medium

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	supplemented with 10% FBS and l-glutamine (1%)

Protocol tips
supplemented with 10% FBS and l-glutamine (1%)

Publication protocol

Human medulloblastoma cell lines Daoy (from a 4-year old male) and UW228 (from a 9-year old female) [27,28] were grown in DMEM media, supplemented with 10% FBS and l-glutamine (1%) at 37°, 5% CO2. Cells were washed 3 times in ice-cold PBS and then collected, using a cell scraper, in lysis buffer containing 10 mM Tris-HCl, pH 7.5, 150 mM NaCl, 1% v/v Nonidet P-40 and protease inhibitor cocktail (Roche Diagnostics, Dee Why, Australia). The suspended cells were incubated for 30 min and ultra-sonicated (Branson Sonifier 450, VWR, Mississauga, ON, Canada) at intervals of 15 s for a total of 2 min, with 15 s pause between each treatment. This was followed by centrifugation at 17,000× g for 1 h. All the above steps were performed at 4 °C. Three biological replicates were performed for each analysis.

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