title: Cell energy budget platform for assessment of cell metabolism.
abstract: Changes in bioenergetic parameters report on metabolic rearrangement, dysfunction of major pathways, and regulatory processes within the cell, adaptation to energy stress, or new physiological condition. A combined measurement of oxidative phosphorylation, glycolytic flux, the Krebs cycle activity, ATP levels, and total biomass allows detailed metabolic assessment. We describe a simple methodology for high-throughput multiparametric assessment of cell bioenergetics, called cell energy budget (CEB) platform, and demonstrate its practical use with cell models. The CEB relies on a standard multi-label reader with time-resolved fluorescence capabilities, the lanthanide probe pH-Xtra™ to measure extracellular acidification (ECA) associated with lactate (L-ECA) and combined lactate/CO2 (T-ECA) extrusion, the phosphorescent probe MitoXpress®-Xtra to measure oxygen consumption rate (OCR), the bioluminescent total ATP assay, and absorbance-based total protein assay. This approach can be further extended with the measurement of other cellular parameters, such as NAD(P)H, Ca(2+), mitochondrial pH, membrane potential, and redox state, using the corresponding fluorescent or luminescent probes.
authors: School of Biochemistry and Cell Biology, University College Cork, Cavanagh Pharmacy Building, Office 1.28, College Road, Cork, Ireland, d.papkovsky@ucc.ie.
authors: Dmitri B
authors: DB
authors: Papkovsky
authors: Alexander V
authors: AV
authors: Zhdanov
mesh_terms: D000818:Animals; D004734:Energy Metabolism; D006019:Glycolysis; D057166:High-Throughput Screening Assays / Q000379:methods; D008163:Luminescent Measurements / Q000379:methods; D010085:Oxidative Phosphorylation; D010101:Oxygen Consumption; D016716:PC12 Cells; D051381:Rats