Lamellipod Reconstruction by Three-Dimensional Reflection Interference Contrast Nanoscopy (3D-RICN)

Area of Science:

• Cell biology
• Biophysics
• Microscopy

Background:

• Existing nanometric cell imaging relies on fluorescence, facing limitations from staining and artifacts.
• Reflection interference contrast microscopy (RICM) is label-free but struggles with complex cellular structures like lamellipodia.

Purpose of the Study:

• To develop a novel label-free technique for reconstructing cell shape at nanometric resolution.
• To quantitatively analyze the lamellipodial region of adherent cells, measuring distance and thickness.

Main Methods:

• Introduced 3D reflection interference contrast nanoscopy (3D-RICN).
• Combined multiple illumination wavelengths and aperture angles for data acquisition.
• Validated using atomic force microscopy and quantitative phase imaging on fixed cells.

Main Results:

• 3D-RICN accurately characterizes lamellipodial thickness and substrate-cell distance.
• Demonstrated radial symmetry in lamellipodial thickness for cells on micropatterns.
• Measured substrate-lamellipod distances up to 100 nm.

Conclusions:

• 3D-RICN offers a powerful label-free approach for nanoscale imaging of cell structures.
• The method is applicable to living cells, enabling dynamic studies.
• Advances understanding of cell adhesion and lamellipodial dynamics at the nanoscale.