Interface-Sensitive Epi-Coherent Anti-Stokes Raman Scattering Microscopy for Imaging Cell Adhesion Dynamics

Area of Science:

• Biophysics
• Cell Biology
• Chemical Imaging

Background:

• Understanding cell adhesion is crucial for cell migration and extracellular matrix interactions.
• Conventional microscopy methods lack chemical specificity or require fluorescent labels for interfacial studies.

Purpose of the Study:

• To develop a label-free microscopy technique for studying cell adhesion dynamics at interfaces.
• To investigate the precise visualization of cell-substrate interactions and lipid droplets using epi-CARS.

Main Methods:

• Utilized homodyne epi-coherent anti-Stokes Raman scattering (epi-CARS) microscopy focused at interfaces.
• Incorporated a pinhole in a confocal epi-CARS system to reject out-of-focus reflections.
• Applied the method to study cell-substrate adhesion dynamics in mitotic and post-mitotic cells.

Main Results:

• Epi-CARS demonstrated intrinsic interfacial selectivity with negative contrast at attachment areas, precisely measuring cell-substrate distances.
• The technique provided high contrast for cellular lipid droplets in an inverted microscope configuration.
• Observed adhesion site splitting during mitosis and outward development during postmitotic spreading.
• Characterized distinct adhesion patterns and water layer thicknesses at leading and retracting cell edges post-mitosis.

Conclusions:

• Homodyne epi-CARS microscopy is a powerful label-free tool for interfacial cell biology.
• The method enables precise nanoscale imaging of cell adhesion and lipid dynamics.
• Revealed dynamic changes in cell adhesion during mitosis and spreading with unprecedented detail.