A novel approach to measure mitochondrial respiration in frozen biological samples

EMBO J. 2020 Jul 1;39(13):e104073. doi: 10.15252/embj.2019104073. Epub 2020 May 20.


Respirometry is the gold standard measurement of mitochondrial oxidative function, as it reflects the activity of the electron transport chain complexes working together. However, the requirement for freshly isolated mitochondria hinders the feasibility of respirometry in multi-site clinical studies and retrospective studies. Here, we describe a novel respirometry approach suited for frozen samples by restoring electron transfer components lost during freeze/thaw and correcting for variable permeabilization of mitochondrial membranes. This approach preserves 90-95% of the maximal respiratory capacity in frozen samples and can be applied to isolated mitochondria, permeabilized cells, and tissue homogenates with high sensitivity. We find that primary changes in mitochondrial function, detected in fresh tissue, are preserved in frozen samples years after collection. This approach will enable analysis of the integrated function of mitochondrial Complexes I to IV in one measurement, collected at remote sites or retrospectively in samples residing in tissue biobanks.

Keywords: frozen tissue; methodology; mitochondrial content; mitochondrial uncoupled respiration; oxygen consumption.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cryopreservation*
  • Electron Transport Chain Complex Proteins / metabolism*
  • Male
  • Mice
  • Mitochondria / metabolism*
  • Oxygen Consumption*
  • Zebrafish / metabolism*
  • Zebrafish Proteins / metabolism*


  • Electron Transport Chain Complex Proteins
  • Zebrafish Proteins