Expansion microscopy-based imaging of nuclear structures in cultured cells

Area of Science:

• Cell biology
• Microscopy techniques
• Biochemistry

Background:

• Expansion microscopy (ExM) is a transformative sample preparation technique.
• It enables nanoscale imaging of biological samples by physically expanding them.
• Conventional microscopy often lacks the resolution to visualize fine cellular structures.

Purpose of the Study:

• To detail the X10 expansion microscopy protocol for U2OS cells.
• To demonstrate nanoscale resolution of nuclear structures.
• To present tools for improved sample handling and analysis in ExM.

Main Methods:

• Utilized the X10 expansion microscopy method for tenfold sample expansion.
• Employed concurrent DNA staining for enhanced visualization.
• Developed custom 3D-printed gel cutters and chambered slides to prevent gel drift.
• Analyzed expanded samples using conventional microscopy and Airyscan confocal imaging.

Main Results:

• Achieved tenfold isotropic expansion of U2OS cells.
• Successfully visualized components of nuclear structures at nanoscale resolution.
• Demonstrated the utility of custom tools in stabilizing gels for improved imaging.
• Validated the protocol for detailed subcellular analysis.

Conclusions:

• The X10 expansion microscopy protocol provides high-resolution nanoscale imaging of cellular components.
• Custom-designed tools enhance the reproducibility and ease of ExM sample preparation.
• This technique is valuable for studying the fine details of nuclear organization and other subcellular structures.