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Real-time Analysis of Metabolic Activity Within Lactobacillus acidophilus by Phasor Fluorescence Lifetime Imaging Microscopy of NADH

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Abstract

Nicotinamide adenine dinucleotide (NADH) is an endogenous fluorescent molecule commonly used as a metabolic biomarker. Fluorescence lifetime imaging microscopy (FLIM) is a method in which the fluorescence decay is measured at each pixel of an image. While the fluorescence spectrum of free and protein-bound NADH is very similar, free and protein-bound NADH display very different decay profiles. Therefore, FLIM can provide a way to distinguish free/bound NADH at the level of single bacteria within biological samples. The phasor technique is a graphical method to analyse the entire image and to produce a histogram of pixels with different decay profile. In this study, NADH fluorescence decay profiles within Lactobacillus acidophilus samples treated using different protocols indicated discernible variations. Clear distinctions between fluorescence decay profiles of NADH in samples of artificially heightened metabolic activity in comparison to those of samples lacking an accessible carbon source were obtained.

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Acknowledgments

We appreciate the help of Krystyna Drozdowicz-Tomsia and Macquarie University for the use of the confocal microscope. KT thanks the University of Western Sydney for the Honours scholarship.

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Authors and Affiliations

  1. School of Science and Health, University of Western Sydney, Richmond, NSW, Australia

    Keenan Torno, Belinda K. Wright, Mark R. Jones & Michael Phillips

  2. The Laboratory for Fluorescence Dynamics, Biomedical Engineering Department, University of California, Irvine, California

    Michelle A. Digman & Enrico Gratton

Authors
  1. Keenan Torno
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  2. Belinda K. Wright
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  3. Mark R. Jones
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  4. Michelle A. Digman
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  5. Enrico Gratton
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  6. Michael Phillips
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Correspondence to Michael Phillips.

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Torno, K., Wright, B.K., Jones, M.R. et al. Real-time Analysis of Metabolic Activity Within Lactobacillus acidophilus by Phasor Fluorescence Lifetime Imaging Microscopy of NADH. Curr Microbiol 66, 365–367 (2013). https://doi.org/10.1007/s00284-012-0285-2

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  • DOI: https://doi.org/10.1007/s00284-012-0285-2

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