ATP content was measured by a commercial bioluminescent assay (ATP bioluminescent assay kit, Merck) according to the manufacturer’s instruction and as previously described [27 (link)]. Briefly, ATP was extracted by boiling the samples in a solution containing 100 mM TRIS, 4 mM EDTA, pH 7.75. After centrifugation at 10,000× g for 60 s, samples were diluted at 1:50 in dilution buffer (FLAA, Merck). To obtain bioluminescence measurements with a standard luminometer, 100 μL of supernatant was mixed with 100 μL of luciferin-luciferase solution. The standard curve of ATP was obtained by serial dilution of 2 μM ATP solution.
Mollo N., Esposito M., Aurilia M., Scognamiglio R., Accarino R., Bonfiglio F., Cicatiello R., Charalambous M., Procaccini C., Micillo T., Genesio R., Calì G., Secondo A., Paladino S., Matarese G., Vita G.D., Conti A., Nitsch L, & Izzo A. (2021). Human Trisomic iPSCs from Down Syndrome Fibroblasts Manifest Mitochondrial Alterations Early during Neuronal Differentiation. Biology, 10(7), 609.
Bioluminescence measurements BufferCentrifugation Dilution Edta Luciferase Luciferin Tris
Corresponding Organization : University of Naples Federico II
Other organizations :
Institute for Experimental Endocrinology and Oncology, National Research Council, Fondazione Santa Lucia, Istituti di Ricovero e Cura a Carattere Scientifico
Volume of supernatant and luciferin-luciferase solution at 100 μL each
positive controls
Standard curve of ATP obtained by serial dilution of 2 μM ATP solution
negative controls
None explicitly mentioned
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