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Updated: May 30, 2026

Autofluorescence Imaging to Evaluate Cellular Metabolism
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Autofluorescence Imaging to Evaluate Cellular Metabolism

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Intracellular pH sensing using autofluorescence lifetime microscopy.

Shinya Ogikubo1, Takakazu Nakabayashi, Takashi Adachi

  • 1Research Institute for Electronic Science, Hokkaido University, Sapporo 001-0020, Japan.

The Journal of Physical Chemistry. B
|July 23, 2011
PubMed
Summary

Intracellular pH can be measured using fluorescence lifetime imaging of reduced nicotinamide adenine dinucleotide (NADH), a key metabolic cofactor. NADH

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Area of Science:

  • Biophysics
  • Cellular Metabolism
  • Fluorescence Spectroscopy

Background:

  • Reduced nicotinamide adenine dinucleotide (NADH) is a crucial cofactor in cellular metabolism.
  • Autofluorescence properties of NADH are sensitive to its microenvironment.
  • Intracellular pH is a critical parameter for cellular function.

Purpose of the Study:

  • To investigate the relationship between intracellular pH and NADH fluorescence lifetime.
  • To determine if NADH fluorescence lifetime imaging can be used for label-free pH sensing in cells.
  • To explore the spatial heterogeneity of NADH fluorescence lifetime within cells.

Main Methods:

  • Acquisition of fluorescence lifetime images of NADH in living cells at various intracellular pH values.
  • Comparison of NADH fluorescence lifetime in cellular compartments (nucleus, mitochondria) versus buffer solutions.

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  • Analysis of the correlation between pH and NADH fluorescence lifetime.

    • Main Results:

    • NADH fluorescence lifetime decreases monotonically with increasing intracellular pH.
    • The pH-induced change in NADH fluorescence lifetime is more pronounced within cells than in buffer.
    • Significant spatial variations in NADH fluorescence lifetime were observed, with shorter lifetimes in the nucleus compared to mitochondria.
    • The magnitude of pH-induced lifetime changes was greater in nuclei.

    Conclusions:

    • Fluorescence lifetime imaging of endogenous NADH provides a label-free method for determining intracellular pH.
    • Spatial heterogeneity of NADH fluorescence lifetime within cells is influenced by pH and potentially local electric fields.
    • This technique offers a novel approach for real-time monitoring of cellular pH dynamics.