Fluorescence lifetime imaging for the two-photon microscope: time-domain and frequency-domain methods

Gratton, Enrico; Breusegem, Sophie; Sutin, Jason; Ruan, Qiaoaio; Barry, Nicolas P. E.

doi:10.1117/1.1586704

Cited by 241 publications

(208 citation statements)

References 36 publications

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“…FLIM is built upon time‐resolved detection and can provide a sensitive contrast mechanism to identify the local environment of a fluorophore 52. Using FLIM, it was determined that two distinct lifetime populations are present for AuNT‐BSA hydrogel and BSA hydrogel.…”

Section: Resultsmentioning

confidence: 99%

Regioselective Localization and Tracking of Biomolecules on Single Gold Nanoparticles

et al. 2015

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Selective localization of biomolecules at the hot spots of a plasmonic nanoparticle is an attractive strategy to exploit the light–matter interaction due to the high field concentration. Current approaches for hot spot targeting are time‐consuming and involve prior knowledge of the hot spots. Multiphoton plasmonic lithography is employed to rapidly immobilize bovine serum albumin (BSA) hydrogel at the hot spot tips of a single gold nanotriangle (AuNT). Regioselectivity and quantity control by manipulating the polarization and intensity of the incident laser are also established. Single AuNTs are tracked using dark‐field scattering spectroscopy and scanning electron microscopy to characterize the regioselective process. Fluorescence lifetime measurements further confirm BSA immobilization on the AuNTs. Here, the AuNT‐BSA hydrogel complexes, in conjunction with single‐particle optical monitoring, can act as a framework for understanding light–molecule interactions at the subnanoparticle level and has potential applications in biophotonics, nanomedicine, and life sciences.

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Section: Resultsmentioning

confidence: 99%

Regioselective Localization and Tracking of Biomolecules on Single Gold Nanoparticles

et al. 2015

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“…The geometry of the beam-multiplexer generates a single line of up to 64 foci in the object plane whereby adjacent foci have opposite polarisation and a typical distance of 1.5 μm from each other for a 20× objective lens (NA = 0.95). In addition, the degree of parallelization can be changed from 64 beams down to 32,16,8,4 and also to a single one whereas the power per beam is doubled with each time the number is halved. Thereby the length of the line is reduced while the spacing between adjacent foci remains constant.…”

Section: Fluorescence Imaging Techniquesmentioning

confidence: 99%

“…However, this emission does not precisely follow the excitation but rather shows phase delays and amplitude changes which are determined by the fluorescence decay law of the sample, i.e. the phase of the emission is shifted to later time points as compared to the excitation light, whereas the peak-to-peak height of the modulated emission is decreased relative to that of the modulated excitation [6][7][8]26,28 .…”

Section: Frequency-domain Flimmentioning

confidence: 99%

“…fluorescence lifetime imaging -FLIM, allow a complete, bioscientifically relevant picture of the samples under study 6,8,26,28 .…”

Section: Principle Of Fluorescence Lifetime Imaging (Flim)mentioning

confidence: 99%

“…homodyne and heterodyne frequency-domain FLIM 8 , time-correlated single-photon counting (TCSPC) 9,10 and time-gated techniques 11,12 as well as on their compatibility with current high-end far-field imaging techniques with particular relevance for the biosciences. The comparison criteria refer to standard fluorescence imaging parameters, i.e.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Fluorescence lifetime imaging in biosciences: technologies and applications

Niesner

Gericke

2008

Front. Phys. China

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The biosciences require the development of methods, which allow a non-invasive and rapid investigation of biological systems. In this frame high-end imaging techniques allow an intravital microscopy in real-time, providing information on a molecular basis. Far-field fluorescence imaging techniques are some of the most adequate methods for such investigations. However, there are great differences between the common fluorescence imaging techniques, i.e. wide-field, confocal one-photon and two-photon microscopy, respectively, as far as their applicability in diverse bioscientific research areas is concerned. In the first part of this work, we briefly compare these techniques. Standard methods used in the biosciences, i.e. steady-state techniques based on the analysis of the total fluorescence signal originating from the sample, can successfully be employed in the study of cell, tissue and organ morphology as well as in monitoring the macroscopic tissue function. However, they are mostly inadequate for the quantitative investigation of the cellular function at molecular level. The intrinsic disadvantages of steady-state techniques are counteracted by using time-resolved techniques. Among these the fluorescence lifetime imaging (FLIM) is currently the most common. Different FLIM principles as well as applications of particular relevance for the biosciences, especially for fast intravital studies are discussed in this work.

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Quantifying Cellular Dynamics by Fluorescence Resonance Energy Transfer (FRET) Microscopy

Grecco

Bastiaens

2013

CP Neuroscience

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The cell is a spatially organized system whose function emerges from the complex interaction of molecular components. Such local interaction of nanometer-sized molecules generates patterns that span throughout the cell. Those patterns, in turn, regulate the molecular interactions. Understanding such simultaneous bidirectional causation requires quantifying the spatio-temporal progression of biochemical reactions in the context of a living cell. Due to its ability to resolve micrometer-sized structures, biological microscopy has been instrumental to the discovery and understanding of living systems. Functional fluorescence microscopy allows a cellular dynamic topographic map of proteins to be overlaid with topological information on the causality that determines protein state. Here we describe how Förster/fluorescence resonance energy transfer (FRET) can be used to measure the phosphorylation state of proteins in the context of the cell.

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Fluorescence lifetime imaging for the two-photon microscope: time-domain and frequency-domain methods

Cited by 241 publications

References 36 publications

Regioselective Localization and Tracking of Biomolecules on Single Gold Nanoparticles

Regioselective Localization and Tracking of Biomolecules on Single Gold Nanoparticles

Fluorescence lifetime imaging in biosciences: technologies and applications

Quantifying Cellular Dynamics by Fluorescence Resonance Energy Transfer (FRET) Microscopy

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