Dataset Information

Light-sheet autofluorescence lifetime imaging with a single-photon avalanche diode array.

ABSTRACT:

Significance

Fluorescence lifetime imaging microscopy (FLIM) of the metabolic co-enzyme nicotinamide adenine dinucleotide (phosphate) [NAD(P)H] is a popular method to monitor single-cell metabolism within unperturbed, living 3D systems. However, FLIM of NAD(P)H has not been performed in a light-sheet geometry, which is advantageous for rapid imaging of cells within live 3D samples.

Aim

We aim to design, validate, and demonstrate a proof-of-concept light-sheet system for NAD(P)H FLIM.

Approach

A single-photon avalanche diode camera was integrated into a light-sheet microscope to achieve optical sectioning and limit out-of-focus contributions for NAD(P)H FLIM of single cells.

Results

An NAD(P)H light-sheet FLIM system was built and validated with fluorescence lifetime standards and with time-course imaging of metabolic perturbations in pancreas cancer cells with 10 s integration times. NAD(P)H light-sheet FLIM in vivo was demonstrated with live neutrophil imaging in a larval zebrafish tail wound also with 10 s integration times. Finally, the theoretical and practical imaging speeds for NAD(P)H FLIM were compared across laser scanning and light-sheet geometries, indicating a 30× to 6× acquisition speed advantage for the light sheet compared to the laser scanning geometry.

Conclusions

FLIM of NAD(P)H is feasible in a light-sheet geometry and is attractive for 3D live cell imaging applications, such as monitoring immune cell metabolism and migration within an organism.

SUBMITTER: Samimi K

PROVIDER: S-EPMC10284079 | biostudies-literature | 2023 Jun

REPOSITORIES: biostudies-literature

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Publications

Light-sheet autofluorescence lifetime imaging with a single-photon avalanche diode array.

Samimi Kayvan K Desa Danielle E DE Lin Wei W Weiss Kurt K Li Joe J Huisken Jan J Miskolci Veronika V Huttenlocher Anna A Chacko Jenu V JV Velten Andreas A Rogers Jeremy D JD Eliceiri Kevin W KW Skala Melissa C MC

Journal of biomedical optics 20230621 6

<h4>Significance</h4>Fluorescence lifetime imaging microscopy (FLIM) of the metabolic co-enzyme nicotinamide adenine dinucleotide (phosphate) [NAD(P)H] is a popular method to monitor single-cell metabolism within unperturbed, living 3D systems. However, FLIM of NAD(P)H has not been performed in a light-sheet geometry, which is advantageous for rapid imaging of cells within live 3D samples.<h4>Aim</h4>We aim to design, validate, and demonstrate a proof-of-concept light-sheet system for NAD(P)H FL ...[more]

PMID: 37351197

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Dataset Information

Light-sheet autofluorescence lifetime imaging with a single-photon avalanche diode array.

Significance

Aim

Approach

Results

Conclusions

Publications

Light-sheet autofluorescence lifetime imaging with a single-photon avalanche diode array.

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