Evidence of the bottom stiffness effect on atomic force microscopy-based cell mechanobiology

Area of Science:

• Biophysics
• Cell Mechanics
• Nanotechnology

Background:

• Atomic Force Microscopy (AFM) is crucial for measuring cell nanomechanical properties.
• Current models often assume semi-infinite cell thickness, neglecting substrate influence.
• Previous studies have not experimentally validated the impact of substrate rigidity on cell mechanics.

Purpose of the Study:

• To investigate the influence of substrate rigidity on forces and apparent moduli measured by AFM.
• To determine if substrate stiffness affects cell mechanical property measurements.
• To develop a method to correct for substrate effects in AFM analysis.

Main Methods:

• Designed a force-distance curve experiment using AFM.
• Applied semi-infinite power-law rheological models for initial data fitting.
• Utilized a bottom-effect correction model to re-analyze force-indentation data.

Main Results:

• Apparent modulus increased with force when using standard models, indicating an artifact.
• This artifact disappeared upon applying a bottom-effect correction model.
• Substrate stiffness intrinsically influences measured forces, not the true mechanical properties of the cell.

Conclusions:

• Substrate rigidity significantly impacts AFM measurements of cell mechanics.
• Standard models can lead to artifacts, overestimating cell stiffness.
• A bottom-effect correction is essential for accurate determination of intrinsic cell mechanical properties.