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Proceedings Paper

A new and highly sensitive method of analyzing metabolic activity using FLIM (Conference Presentation)
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Paper Abstract

Traditional assessments of cellular metabolism are often destructive, time consuming and without visual information. Fluorescence lifetime imaging microscopy (FLIM) provides a highly sensitive, non-invasive, and label-free alternative. This study uses FLIM in combination with two-photon microscopy to investigate pharmacological induced metabolic changes of adipocytes via changes in the fluorescence of the metabolic co-factors NADH and FAD. In agreement with recent publications NADH fluorescence suggests the presence of four distinct lifetimes in cell culture and tissue with two unbound and two protein bound states which show different responses to treatment with metabolic modifiers. We evaluated the effects on NADH fluorescence lifetime after systematic manipulations to change the balance between oxidative and glycolytic metabolism using five pharmacological reagents - Oligomycin, 2-DG, FCCP, Rotenone, and Glucose - which interact with different parts of the metabolic pathway. We established several ratios between the four distinct lifetimes of NADH after treatment and compared the results to oxygen consumption rate and extracellular acidification rate. We demonstrated, for the first time, a correlation between the two unbound fluorescence lifetimes components and glycolytic and oxidative metabolic activity with a significant higher sensitivity compared to the commonly used free-to-bound ratio of NADH. Analyzing all four lifetime components of NADH has the potential to become a powerful tool to evaluate metabolic activity of adipocytes with subcellular resolution.

Paper Details

Date Published: 24 April 2017
PDF: 1 pages
Proc. SPIE 10069, Multiphoton Microscopy in the Biomedical Sciences XVII, 100691O (24 April 2017); doi: 10.1117/12.2252339
Show Author Affiliations
Michael Evers, Massachusetts General Hospital (United States)
Nunciada Salma, Massachusetts General Hospital (United States)
Reginald Birngruber, Univ. zu Lübeck (Germany)
Conor L. Evans, Harvard Medical School (United States)
Dieter Manstein, Harvard Medical School (United States)


Published in SPIE Proceedings Vol. 10069:
Multiphoton Microscopy in the Biomedical Sciences XVII
Ammasi Periasamy; Peter T. C. So; Karsten König; Xiaoliang S. Xie, Editor(s)

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