4.6. ATP Analysis

Following the culture period, cells were transferred to 2.5 mM glucose in SAB buffer for 2 h, followed by transfer to either 2.5 mM glucose SAB buffer or 16.7 mM glucose SAB buffer for 1 h. Cells were washed with PBS, harvested by trypsinisation and pelleted by centrifugation. The cells were lysed in 150 μL 1M perchloric acid on ice to precipitate cellular proteins. Lysate was centrifuged at 20,000× g for 10 min, after which 150 μL supernatant was transferred to a new tube with 150 μL 1M KOH. ATP was quantified using the ATP assay kit (Life Technologies, Carlsbad, CA, USA).