Determination of ATP, ADP, and AMP Levels by Reversed-Phase High-Performance Liquid Chromatography in Cultured Cells

Methods Mol Biol. 2019:1925:223-232. doi: 10.1007/978-1-4939-9018-4_19.

Abstract

Cytoplasmic and mitochondrial Ca2+ signals couple cellular ATP production to activity-related energy demand. In order to accurately determine the bioenergetic effect of Ca2+ signals, cellular energy charge, i.e., the compound ratio of the phosphorylated adenine nucleotides AMP, ADP, and ATP, should be estimated. Reversed-phase high-performance liquid chromatography (RP-HPLC) allows the rapid separation and quantitation of these molecules. Here we describe a protocol applied in our laboratories to quantify ATP, ADP, and AMP nucleotides in cellular extracts.

Keywords: ATP/ADP ratio; Ca2+ signal; Energy charge; Glycolysis; Oxidative phosphorylation; Reversed-phase high-performance liquid chromatography (RP-HPLC).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Diphosphate / analysis*
  • Adenosine Monophosphate / analysis*
  • Adenosine Triphosphate / analysis*
  • Breast Neoplasms / chemistry
  • Cell Line, Tumor
  • Chromatography, High Pressure Liquid / methods*
  • Chromatography, Reverse-Phase / methods*
  • Female
  • Humans
  • MCF-7 Cells

Substances

  • Adenosine Monophosphate
  • Adenosine Diphosphate
  • Adenosine Triphosphate