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Fluorescence Lifetime Macro Imager for Biomedical Applications
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Light-sheet autofluorescence lifetime imaging with a single-photon avalanche diode array.

Kayvan Samimi1, Danielle E Desa1, Wei Lin2

  • 1Morgridge Institute for Research, Madison, Wisconsin, United States.

Journal of Biomedical Optics
|June 23, 2023
PubMed
Summary
This summary is machine-generated.

We developed a novel light-sheet fluorescence lifetime imaging microscopy (FLIM) system for monitoring cellular metabolism using nicotinamide adenine dinucleotide (phosphate) [NAD(P)H]. This advanced technique offers a significant speed advantage for live 3D cell imaging applications.

Keywords:
autofluorescencefluorescence lifetime imaging microscopylight-sheet microscopynicotinamide adenine dinucleotide (phosphate)single-photon avalanche diode

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Area of Science:

  • Biophysics
  • Cellular Metabolism
  • Microscopy

Background:

  • Fluorescence lifetime imaging microscopy (FLIM) of nicotinamide adenine dinucleotide (phosphate) [NAD(P)H] is crucial for studying single-cell metabolism in living 3D systems.
  • Current FLIM methods for NAD(P)H lack the speed and optical sectioning capabilities offered by light-sheet microscopy, particularly for rapid imaging of live 3D samples.

Purpose of the Study:

  • To design, validate, and demonstrate a proof-of-concept light-sheet system for NAD(P)H FLIM.
  • To enable faster and more efficient 3D live cell imaging of metabolic processes.

Main Methods:

  • Integration of a single-photon avalanche diode camera into a light-sheet microscope.
  • Optical sectioning to minimize out-of-focus light contributions.
  • Validation using fluorescence lifetime standards and metabolic perturbation studies in pancreas cancer cells.

Main Results:

  • Successful construction and validation of the NAD(P)H light-sheet FLIM system.
  • Demonstration of 10-second integration time imaging of metabolic changes in cancer cells and in vivo imaging of neutrophils in zebrafish.
  • Quantified a significant speed advantage (up to X-fold) for light-sheet geometry compared to laser scanning for NAD(P)H FLIM.

Conclusions:

  • Light-sheet geometry is feasible and highly advantageous for NAD(P)H FLIM.
  • This technology is attractive for 3D live cell imaging, including monitoring immune cell metabolism and migration within organisms.