Monitoring changes in endogenous fluorophores through quantitative FLIM imaging in live cells

Vinod Jyothikumar; Yuansheng Sun; Ammasi Periasamy

doi:10.1117/12.923666

9 February 2012 Monitoring changes in endogenous fluorophores through quantitative FLIM imaging in live cells

Vinod Jyothikumar, Yuansheng Sun, Ammasi Periasamy

Proceedings Volume 8226, Multiphoton Microscopy in the Biomedical Sciences XII; 822644 (2012) https://doi.org/10.1117/12.923666
Event: SPIE BiOS, 2012, San Francisco, California, United States

Abstract

Changes in energy metabolism, mitochondrial functions and of reactive oxygen species are often supposed to induce alterations in cellular activity. The major intracellular endogenous fluorophores are reduced nicotinamide adenine dinucleotide (NADH) and dinucleotide phosphate (NADPH), riboflavin's, and tryptophan present inside biological tissue and they can be used to image tissue architecture without any exogenous probe. Their fluorescence can be excited by multi-photon microscopy using NIR laser wavelengths [1,2,5,6]. Using FLIM imaging the lifetime of tryptophan and NADH were monitored in cells with and without addition of glucose in the medium. The lifetime data were collected and further using the ANOVA (refer table 2) of the lifetime of free NADH, bound NADH and Tryptophan, we found on applying the null hypothesis for the P-value > 0.05, there is a significant difference between the lifetimes of bound NADH and Tryptophan from control to glucose treated cells, however, free NADH, shown to be not significant change between the control and glucose treated cells. The tryptophan puzzle comes closer and closer to a solution. The ultimate evidence supporting the existence of FRET between Tryptophan and NADH in live cells slowly could come from lifetime measurements of tryptophan in proteins and bound NADH within live cells.

Citation Download Citation

Vinod Jyothikumar, Yuansheng Sun, and Ammasi Periasamy "Monitoring changes in endogenous fluorophores through quantitative FLIM imaging in live cells", Proc. SPIE 8226, Multiphoton Microscopy in the Biomedical Sciences XII, 822644 (9 February 2012); https://doi.org/10.1117/12.923666

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KEYWORDS

Luminescence

Fluorescence lifetime imaging

Tissues

Glucose

Cancer

Fluorescence resonance energy transfer

Mode conditioning cables

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