Multiplane 2.5D microscopy for high-throughput high-resolution tissue imaging

Area of Science:

• Biomedical Imaging
• Microscopy Techniques
• Tissue Analysis

Background:

• High-throughput fluorescence imaging is crucial for biomedical applications, especially high-resolution volumetric tissue analysis.
• Current methods can be time-consuming, limiting the scope of tissue studies.

Purpose of the Study:

• To develop and characterize an advanced imaging strategy for fast, high-resolution volumetric tissue analysis.
• To combine multiplane microscopy with extended depth-of-field (EDOF) microscopy.

Main Methods:

• Utilized 2.5D microscopy, an EDOF approach, integrated with a quad-plane image splitter.
• Enabled simultaneous capture of four focal volumes for volumetric imaging of 16–20 μm thick mouse and human tissues.
• Demonstrated compatibility with multicolor imaging and nucleus segmentation.

Main Results:

• Achieved a 25-fold reduction in image acquisition time compared to conventional z-scanning widefield microscopy.
• A 2 mm x 2 mm x 16 μm volume was imaged in 4.7 minutes, down from approximately 2 hours.
• Successful application to nucleus segmentation for downstream analysis was demonstrated.

Conclusions:

• The developed imaging technique offers a significant improvement in volumetric imaging speed for tissue analysis.
• Provides a promising tool for researchers with minimal compromise in spatial resolution and sensitivity.
• Facilitates faster and more efficient high-resolution volumetric tissue studies.