4.4. Glucose Transport Assay

The glucose transport assay was performed on differentiated C2C12 myotubes. The glucose uptake activity was detected using 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-amino]-2-deoxy-D-glucose (2-NBDG; Life Technologies, Grand Island, NY, USA), a fluorescent D-glucose analog, as previously described [41]. Briefly, myotubes were pre-cultured in Krebs Henseleit buffer (pH 7.4 with 2% BSA) containing 0.05% glucose at 37℃ for 30 min. Myotubes were then treated with 2-NBDG (200 μM) for 30 min in glucose-free Krebs Henseleit buffer with 2% BSA. The fluorescence intensities of 2-NBDG uptake were measured by a microplate fluorometer (Beckman Coulter, Brea, CA, USA) with λex = 485 nm and λem = 535 nm.