Simultaneous Live Imaging of Multiple Insect Embryos in Sample Chamber-Based Light Sheet Fluorescence Microscopes

Area of Science:

• Developmental Biology
• Microscopy
• Genetics

Background:

• Light sheet-based fluorescence microscopy is crucial for studying biological processes across scales.
• Sample chamber setups with rotation are ideal for developmental biology, enabling 3D imaging of specimens like Drosophila and Tribolium.
• Current live imaging protocols often focus on single embryos, limiting throughput and introducing ambient variance in comparative studies.

Purpose of the Study:

• To develop an experimental framework for simultaneous live imaging in sample chamber-based setups.
• To increase throughput and ensure consistent ambient conditions for multiple specimens.
• To facilitate comparative studies and quantitative analyses in developmental biology.

Main Methods:

• A calibration guideline for light sheet fluorescence microscopes was established.
• A novel mounting method for multiple embryos compatible with sample rotation was proposed.
• Exemplary 3D live imaging datasets were generated for Drosophila and Tribolium.

Main Results:

• The developed framework allows for simultaneous live imaging of multiple specimens.
• The protocol addresses ambient variance inherent in sequential live imaging.
• Increased throughput was achieved, beneficial for limited microscope access.

Conclusions:

• The proposed protocol enhances throughput and reduces ambient variance for comparative live imaging studies.
• This method is particularly valuable for quantitative analyses and characterizing phenotypes, such as those from knockout experiments.
• The mounting technique is adaptable to other insect species and model organisms like zebrafish.