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Using Light Sheet Fluorescence Microscopy to Image Zebrafish Eye Development

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Published on: April 10, 2016

Multiview deconvolution approximation multiphoton microscopy of tissues and zebrafish larvae.

Dimitrios Kapsokalyvas^1,2, Rodrigo Rosas³, Rob W A Janssen³

¹Department of Genetics and Cell Biology, Faculty of Health, Medicine and Life Sciences (FHML), Maastricht University, Maastricht, the Netherlands. d.kapsokalyvas@maastrichtuniversity.nl.

Scientific Reports

|May 13, 2021

Summary

Multiview imaging enhances multiphoton microscopy for clearer 3D visualization of tissues and organisms. This technique improves resolution and suppresses artifacts, aiding in the accurate quantification of small structures.

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Area of Science:

Biomedical Imaging
Optical Microscopy
3D Reconstruction

Background:

Tomographic microscopy techniques like confocal, multiphoton, and light sheet microscopy are essential for imaging thick tissues and small organisms.
These techniques often face challenges with anisotropic resolution and limited penetration depth.
Multiview microscopy, involving imaging from multiple angles and 3D reconstruction, was previously applied to light sheet microscopy to overcome these limitations.

Purpose of the Study:

To adapt and apply Multiview imaging methodology to multiphoton microscopy.
To achieve isotropic resolution and visualize entire samples with improved clarity.
To suppress interference artifacts for better visualization of biological structures.

Main Methods:

Applied Multiview imaging to multiphoton microscopy, utilizing both fluorescence and second harmonic signals.
Acquired images of samples from multiple angles.
Reconstructed 3D images from the acquired data.

Main Results:

Achieved isotropic resolution in 3D imaging.
Enabled visualization of the entire sample, including collagen fibrils and myofibrils.
Successfully suppressed interference artifacts, leading to clear imaging.
Demonstrated applicability to various scanning microscopy techniques without modifications.

Conclusions:

Multiview imaging combined with multiphoton microscopy significantly improves 3D imaging capabilities for tissues and small organisms.
The method offers at least a threefold axial resolution improvement, crucial for accurate quantification of 3D structures.
This technique is versatile, applicable to both label-free and stained samples, including heart tissue and zebrafish larvae.