Multiphoton fluorescence lifetime imaging microscopy reveals free-to-bound NADH ratio changes associated with metabolic inhibition

Krystyna Drozdowicz-Tomsia*, Ayad G. Anwer, Michael A. Cahill, Kaiser N. Madlum, Amel M. Maki, Mark S. Baker, Ewa M. Goldys

*Corresponding author for this work

Research output: Contribution to journalArticle

32 Citations (Scopus)
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Measurement of endogenous free and bound NAD(P)H relative concentrations in living cells is a useful method for monitoring aspects of cellular metabolism, because the NADH/NAD\+ reduction-oxidation pair is crucial for electron transfer through the mitochondrial electron transport chain. Variations of free and bound NAD(P)H ratio are also implicated in cellular bioenergetic and biosynthetic metabolic changes accompanying cancer. This study uses two-photon fluorescence lifetime imaging microscopy (FLIM) to investigate metabolic changes in MCF10A premalignant breast cancer cells treated with a range of glycolysis inhibitors: Namely, 2 deoxy-D-glucose, oxythiamine, lonidamine, and 4-(chloromethyl) benzoyl chloride, as well as the mitochondrial membrane uncoupling agent carbonyl cyanide m-chlorophenylhydrazone. Through systematic analysis of FLIM data from control and treated cancer cells, we observed that all glycolytic inhibitors apart from lonidamine had a slightly decreased metabolic rate and that the presence of serum in the culture medium generally marginally protected cells from the effect of inhibitors. Direct production of glycolytic L-lactate was also measured in both treated and control cells. The combination of these two techniques gave valuable insights into cell metabolism and indicated that FLIM was more sensitive than traditional biochemical methods, as it directly measured metabolic changes within cells as compared to quantification of lactate secreted by metabolically active cells.

Original languageEnglish
Article number086016
Pages (from-to)1-13
Number of pages13
JournalJournal of Biomedical Optics
Issue number8
Publication statusPublished - Aug 2014

Bibliographical note

Copyright the Author(s) 2014. Version archived for private and non-commercial use with the permission of the author/s and according to publisher conditions. For further rights please contact the publisher.


  • optics
  • photonics
  • light
  • lasers
  • fluorescence
  • fluorescence lifetime imaging

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