Integrated Array Tomography for 3D Correlative Light and Electron Microscopy.

Area of Science:

• Biophysics
• Cell Biology
• Microscopy

Background:

• Large-scale volume electron microscopy (EM) is crucial for biological systems but faces challenges with slow acquisition rates and lack of targeted information.
• Existing methods often generate excessive data, complicating analysis and limiting biological insights.

Purpose of the Study:

• To develop and validate a 3D image acquisition and reconstruction pipeline that integrates fluorescence microscopy (FM) and scanning electron microscopy (SEM).
• To overcome the limitations of slow acquisition and non-specific data in large-scale EM by enabling targeted imaging within biological samples.

Main Methods:

• Integration of a widefield fluorescence microscope within a scanning electron microscope.
• Utilizing large field-of-view FM images to guide subsequent high-resolution EM acquisition of regions of interest.
• Employing cathodoluminescent markers for precise overlay and correlation between EM and FM datasets.

Main Results:

• Demonstrated proof-of-concept on immunolabelled serial tissue sections.
• Significantly expedited total acquisition times by focusing EM on targeted biological regions.
• Reduced data burden by tens or hundreds of gigabytes through selective data collection.

Conclusions:

• The integrated correlative light and electron microscopy (CLEM) workflow effectively addresses key limitations of standalone large-scale EM.
• This approach enhances efficiency and biological relevance in high-resolution 3D imaging of tissues.
• Facilitates faster, more targeted, and data-efficient volumetric analysis in biological research.