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Visualizing Intracellular SNARE Trafficking by Fluorescence Lifetime Imaging Microscopy
Published on: December 29, 2017
Imaging intracellular signaling using two-photon fluorescent lifetime imaging microscopy.
Cold Spring Harbor Protocols
|November 3, 2012
Summary
Förster resonance energy transfer (FRET) sensors and imaging enable visualization of intracellular signaling dynamics. Two-photon fluorescence lifetime imaging microscopy (2pFLIM) offers powerful quantitative imaging in thick tissues.
Area of Science:
- Cellular and Molecular Biology
- Biophysics
- Microscopy Techniques
Background:
- Förster resonance energy transfer (FRET) sensors and imaging techniques allow visualization of intracellular signaling dynamics.
- High spatiotemporal resolution is crucial for monitoring rapid cellular events.
- Subcellular compartments and thick tissues present imaging challenges.
Purpose of the Study:
- To provide practical guidelines for quantitative imaging of intracellular signaling.
- To highlight the utility of two-photon fluorescence lifetime imaging microscopy (2pFLIM) for this purpose.
- To enable researchers to effectively utilize 2pFLIM for studying cellular signaling.
Main Methods:
- Utilizing Förster resonance energy transfer (FRET) sensors for molecular event detection.
- Employing fluorescence lifetime imaging microscopy (FLIM).
- Combining FLIM with two-photon laser-scanning microscopy (2pFLIM) for enhanced tissue penetration and resolution.
Main Results:
- 2pFLIM enables high spatiotemporal resolution imaging of intracellular signaling.
- The technique is effective for monitoring events in small subcellular compartments.
- Quantitative imaging in thick tissue samples is feasible with 2pFLIM.
Conclusions:
- 2pFLIM is a powerful tool for quantitative imaging of intracellular signaling dynamics.
- This method facilitates the study of signaling in challenging biological samples.
- Practical guidelines are provided to aid researchers in applying 2pFLIM.
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