Calculation of ATP production rates using the Seahorse XF Analyzer

Brandon R. Desousa, Kristen K.O. Kim, Anthony E. Jones, Andréa B. Ball, Wei Y. Hsieh, Pamela Swain, Danielle H. Morrow, Alexandra J. Brownstein, David A. Ferrick, Orian S. Shirihai, Andrew Neilson, David A. Nathanson, George W. Rogers, Brian P. Dranka, Anne N. Murphy, Charles Affourtit, Steven J. Bensinger, Linsey Stiles, Natalia Romero, Ajit S. Divakaruni

Research output: Contribution to journalArticlepeer-review

17 Scopus citations

Abstract

Oxidative phosphorylation and glycolysis are the dominant ATP-generating pathways in mammalian metabolism. The balance between these two pathways is often shifted to execute cell-specific functions in response to stimuli that promote activation, proliferation, or differentiation. However, measurement of these metabolic switches has remained mostly qualitative, making it difficult to discriminate between healthy, physiological changes in energy transduction or compensatory responses due to metabolic dysfunction. We therefore present a broadly applicable method to calculate ATP production rates from oxidative phosphorylation and glycolysis using Seahorse XF Analyzer data and empirical conversion factors. We quantify the bioenergetic changes observed during macrophage polarization as well as cancer cell adaptation to in vitro culture conditions. Additionally, we detect substantive changes in ATP utilization upon neuronal depolarization and T cell receptor activation that are not evident from steady-state ATP measurements. This method generates a single readout that allows the direct comparison of ATP produced from oxidative phosphorylation and glycolysis in live cells. Additionally, the manuscript provides a framework for tailoring the calculations to specific cell systems or experimental conditions.

Original languageEnglish
Article numbere56380
JournalEMBO Reports
Volume24
Issue number10
DOIs
StatePublished - 9 Oct 2023
Externally publishedYes

Keywords

  • ATP
  • ECAR
  • Seahorse XF Analyzer
  • glycolysis
  • oxidative phosphorylation

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Genetics

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