Luciferin potassium salt

Luciferin potassium salt (Sigma-Aldrich, Zwijndrecht, the Netherlands) was added to cells at a final concentration of 0.1 mM for 30 min at 37°C. The luciferase activity was measured with a LumiStar Optima luminescence counter (BMG Lab Tech, Offenburg, Germany).

1,1’-dioctadecyl-3,3,3’,3’–tetramethylind-odicarbocyanine, 4-chlorobenzenesulfonate salt (DiD) and 1,1’-dioctadecyl-3,3,3’,3’-tetramethylindotricarbocyanine iodide (DiR) were purchased from Fanbo Biochemical Co., Ltd. (Beijing, China). D-luciferase ((S)-2-(6-Hydroxy-2-benzothiazolyl)-2-thiazoline-4-carboxylic acid potassium salt, 4,5-Dihydro-2-(6-hydroxy-2-benzothiazolyl)-4-thiazolecarboxylic acid potassium salt, Firefly luciferin potassium salt) was purchased from Sigma-Aldrich. (St. Louis, USA) Cholesterol was purchased from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China). Egg yolk phospholipid (EPC) was provided by A.V.T Pharmaceutical Co., Ltd. (Shanghai, China).

Pancreatic acinar cells (1 × 10⁶/condition) were pretreated with antioxidant for 30 min and washed with buffer A (25 mM Tris-HCl, 10 mM KH₂PO₄, 150 mM KCl, 5 mM MgCl₂, 0.1% BSA, pH 7.8). The cells were then covered and permeabilized with 200 µL 1× ATP-releasing reagent in buffer A (Sigma-Aldrich) per well for 2 min. Twenty microlitres of each supernatant was then transferred to a white plate and the measurement protocol started immediately using a POLARstar Omega Plate Reader (BMG Labtech, Germany). Eighty microlitres of mastermix, consisting of 0.3 mM luciferin potassium salt and luciferase (Sigma-Aldrich), was injected per well and the luminescence emission recorded for 15 min. Addition of the ATP synthase inhibitor oligomycin was used to show maximal blockade of mitochondrial respiration. The chemiluminescence intensity was normalised to negative controls for each mouse/run.