Q700 ice water bath sonicator

Powdered GC muscles (10–20 mg per assay) were sonicated in the appropriate assay buffer (Q700 ice-water bath sonicator, QSONICA) for six cycles of amplitude 35 for 30 sec followed by 1 min on ice, centrifuged at 10,000 × g for 10 min at 4 °C and the lysates were subjected to the following assays: measurement of the metabolites and citrate synthase (CS) activity. Lactate Colorimetric Assay (Biovision, #K607-100), Glycogen Colorimetric Assay (Biovision, #K648-100), and Citrate Synthase Activity Colorimetric Assay (Biovision, #K318-100) were performed according to the manufacturers’ instructions.

Powdered GC muscles (10-15 mg) were sonicated in 3% trichloroacetic acid (TCA)/PBS (Q700 ice-water bath sonicator, QSONICA) for six cycles at amplitude 35 for 30 s followed by 1 min on ice and centrifuged at 12,000×g for 5 min at 4 °C. The supernatant was transferred into a new ice-cold tube, diluted with an equal volume of ddH₂O, and pH was adjusted to neutral with 1 M KOH. The amounts of ATP, ADP and AMP in each sample were measured by UV-HPLC in the Analytical Pharmacology Core at City of Hope (Duarte, CA). The pellet after centrifugation was dissolved in 5% sodium dodecyl sulfate (SDS)/ 0.1 N NaOH, and protein concentration was measured by BCA assay (Thermo Fisher Scientific).