Correlation of live-cell imaging with volume scanning electron microscopy

Area of Science:

• Cell Biology
• Microscopy

Background:

• Live-cell imaging is a fundamental technique in life sciences.
• Correlative microscopy combines live-cell imaging with high-resolution techniques like scanning electron microscopy (SEM).

Purpose of the Study:

• To describe two adaptable live-cell imaging setups compatible with volume SEM.
• To present resin embedding methods for correlative light and electron microscopy.

Main Methods:

• Utilized cell culture dishes with gridded glass supports for versatile light microscopy.
• Employed a flow-chamber setup using Ibidi μ-slides for live-cell imaging.
• Applied serial blockface-SEM and focused ion beam-SEM for high-resolution imaging.
• Described en-bloc embedding and thin-layer plastification for resin embedding.

Main Results:

• Both described live-cell imaging strategies can be successfully integrated with volume SEM.
• Thin-layer plastification is optimal for focused ion beam-SEM, especially for substrate interaction studies.
• En-bloc embedding offers broad applicability for both volume SEM techniques.
• Strategies for relocating cells of interest were discussed for different embedding and microscopy approaches.

Conclusions:

• The presented live-cell imaging setups and embedding techniques facilitate correlative studies.
• These methods enhance the ability to study cellular dynamics and ultrastructure in conjunction with light microscopy.