Two-dimensional fluorescence lifetime imaging for in-vitro and in-vivo application

1998 ◽  
Author(s):  
Paul M. W. French ◽  
Mark J. Dayel ◽  
Keith Dowling ◽  
Sam C. W. Hyde ◽  
M. J. Lever ◽  
...  
Keyword(s):  
Fluorescence Lifetime ◽  
Fluorescence Lifetime Imaging ◽  
Two Dimensional ◽  
Lifetime Imaging

scholarly journals In vivo fluorescence lifetime imaging captures metabolic changes in macrophages during wound responses in zebrafish

2020 ◽  
Author(s):  
Veronika Miskolci ◽  
Kelsey E Tweed ◽  
Michael R Lasarev ◽  
Emily C Britt ◽  
Courtney E McDougal ◽  
...  
Keyword(s):  
Fluorescence Lifetime ◽  
Metabolic Regulation ◽  
Immune Cell ◽  
Sterile Inflammation ◽  
Fluorescence Lifetime Imaging ◽  
Redox Ratio ◽  
Lifetime Imaging ◽  
Intracellular Metabolism

AbstractThe effector functions of macrophages across the spectrum of activation states in vitro are linked to profound metabolic rewiring. However, the metabolism of macrophages remains poorly characterized in vivo. To assess changes in the intracellular metabolism of macrophages in their native inflammatory microenvironment, we employed two-photon fluorescence lifetime imaging microscopy (FLIM) of metabolic coenzymes NAD(P)H and FAD. We found that pro-inflammatory activation of macrophages in vivo was associated with a decrease in the optical redox ratio [NAD(P)H/(NAD(P)H+FAD)] relative to a pro-resolving population during both infected and sterile inflammation. FLIM also resolved temporal changes in the optical redox ratio and lifetime variables of NAD(P)H in macrophages over the course of sterile inflammation. Collectively, we show that non-invasive and label-free imaging of autofluorescent metabolic coenzymes is sensitive to dynamic changes in macrophage activation in interstitial tissues. This imaging-based approach has broad applications in immunometabolism by probing in real time the temporal and spatial metabolic regulation of immune cell function in a live organism.SignificanceMetabolic regulation of macrophage effector functions has recently emerged as a key concept in immune cell biology. Studies rely on in vitro and ex vivo approaches to study macrophage metabolism, however the high plasticity of these cells suggest that removal from their native microenvironment may induce changes in their intracellular metabolism. Here, we show that fluorescence lifetime imaging microscopy of metabolic coenzymes captures dynamic changes in the metabolic activity of macrophages while maintaining them in their endogenous microenvironment. This approach also resolves variations on a single-cell level, in contrast to bulk measurements provided by traditional biochemical assays, making it a potentially valuable tool in the field of immunometabolism.

scholarly journals In vitro and in vivo NIR Fluorescence Lifetime Imaging with a time-gated SPAD camera

2021 ◽  
Author(s):  
Jason T. Smith ◽  
Alena Rudkouskaya ◽  
Shan Gao ◽  
Juhi M. Gupta ◽  
Arin Ulku ◽  
...  
Keyword(s):  
Fluorescence Lifetime ◽  
Near Infrared ◽  
Single Photon ◽  
In Vivo Studies ◽  
Fluorescence Lifetime Imaging ◽  
Iccd Camera ◽  
Lifetime Imaging ◽  
Nir Fluorescence

Near-infrared (NIR) fluorescence lifetime imaging (FLI) provides a unique contrast mechanism to monitor biological parameters and molecular events in vivo. Single-photon avalanche photodiode (SPAD) cameras have been recently demonstrated in FLI microscopy (FLIM) applications, but their suitability for in vivo macroscopic FLI (MFLI) in deep tissues remains to be demonstrated. Herein, we report in vivo NIR MFLI measurement with SwissSPAD2, a large time-gated SPAD camera. We first benchmark its performance in well-controlled in vitro experiments, ranging from monitoring environmental effects on fluorescence lifetime, to quantifying Förster Resonant Energy Transfer (FRET) between dyes. Next, we use it for in vivo studies of target-drug engagement in live and intact tumor xenografts using FRET. Information obtained with SwissSPAD2 was successfully compared to that obtained with a gated-ICCD camera, using two different approaches. Our results demonstrate that SPAD cameras offer a powerful technology for in vivo preclinical applications in the NIR window.

scholarly journals In Vivo Fluorescence Lifetime Imaging for Monitoring the Efficacy of the Cancer Treatment

2014 ◽  
Vol 20 (13) ◽  
pp. 3531-3539 ◽  
Author(s):  
Yasaman Ardeshirpour ◽  
Victor Chernomordik ◽  
Moinuddin Hassan ◽  
Rafal Zielinski ◽  
Jacek Capala ◽  
...  
Keyword(s):  
Cancer Treatment ◽  
Fluorescence Lifetime ◽  
Fluorescence Lifetime Imaging ◽  
Lifetime Imaging ◽  
In Vivo Fluorescence

scholarly journals Coarse-grained modeling of mitochondrial metabolism enables subcellular flux inference from fluorescence lifetime imaging microscopy

2020 ◽  
Author(s):  
Xingbo Yang ◽  
Daniel J. Needleman
Keyword(s):  
Fluorescence Lifetime ◽  
Metabolic Flux ◽  
Living Cells ◽  
Coarse Grained ◽  
Fluorescence Lifetime Imaging Microscopy ◽  
Fluorescence Lifetime Imaging ◽  
Lifetime Imaging ◽  
Metabolic Fluxes ◽  
Subcellular Resolution

AbstractMitochondria are central to metabolism and their dysfunctions are associated with many diseases1–9. Metabolic flux, the rate of turnover of molecules through a metabolic pathway, is one of the most important quantities in metabolism, but it remains a challenge to measure spatiotemporal variations in mitochondrial metabolic fluxes in living cells. Fluorescence lifetime imaging microscopy (FLIM) of NADH is a label-free technique that is widely used to characterize the metabolic state of mitochondria in vivo10–18. However, the utility of this technique has been limited by the inability to relate FLIM measurement to the underlying metabolic activities in mitochondria. Here we show that, if properly interpreted, FLIM of NADH can be used to quantitatively measure the flux through a major mitochondrial metabolic pathway, the electron transport chain (ETC), in vivo with subcellular resolution. This result is based on the use of a coarse-grained NADH redox model, which we test in mouse oocytes subject to a wide variety of perturbations by comparing predicted fluxes to direct biochemical measurements and by self-consistency criterion. Using this method, we discovered a subcellular spatial gradient of mitochondrial metabolic flux in mouse oocytes. We showed that this subcellular variation in mitochondrial flux correlates with a corresponding subcellular variation in mitochondrial membrane potential. The developed model, and the resulting procedure for analyzing FLIM of NADH, are valid under nearly all circumstances of biological interest. Thus, this approach is a general procedure to measure metabolic fluxes dynamically in living cells, with subcellular resolution.

Towards in-vivo assessment of fluorescence lifetime: imaging using time-gated intensified CCD camera

2018 ◽  
Vol 38 (4) ◽  
pp. 966-974 ◽  
Author(s):  
Piotr Sawosz ◽  
Stanislaw Wojtkiewicz ◽  
Michal Kacprzak ◽  
Elzbieta Zieminska ◽  
Magdalena Morawiec ◽  
...  
Keyword(s):  
Fluorescence Lifetime ◽  
Ccd Camera ◽  
Fluorescence Lifetime Imaging ◽  
Lifetime Imaging ◽  
In Vivo Assessment

Using in vivo multiphoton fluorescence lifetime imaging to unravel disease-specific changes in the liver redox state

2020 ◽  
Vol 8 (3) ◽  
pp. 034003
Author(s):  
Deborah S Barkauskas ◽  
Gregory Medley ◽  
Xiaowen Liang ◽  
Yousuf H Mohammed ◽  
Camilla A Thorling ◽  
...  
Keyword(s):  
Fluorescence Lifetime ◽  
Redox State ◽  
Fluorescence Lifetime Imaging ◽  
Lifetime Imaging ◽  
Disease Specific

scholarly journals Fluorescence lifetime imaging of upper gastrointestinal pH in vivo with a lanthanide based near-infrared τ probe

2019 ◽  
Vol 10 (15) ◽  
pp. 4227-4235 ◽  
Author(s):  
Yingying Ning ◽  
Shengming Cheng ◽  
Jing-Xiang Wang ◽  
Yi-Wei Liu ◽  
Wei Feng ◽  
...  
Keyword(s):  
Fluorescence Lifetime ◽  
Near Infrared ◽  
Lanthanide Complex ◽  
Fluorescence Lifetime Imaging ◽  
Ph Responsive ◽  
Gastrointestinal Ph ◽  
Lifetime Imaging ◽  
Upper Gastrointestinal

Lanthanide complex was successfully applied in the design of pH-responsive NIR τ probe for quantitative in vivo imaging.

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