Conditions to minimize soft single biomolecule deformation when imaging with atomic force microscopy

Area of Science:

• Biophysics
• Materials Science
• Nanotechnology

Background:

• Atomic Force Microscopy (AFM) is crucial for imaging soft biomolecules.
• Biomolecule deformation during AFM imaging can lead to inaccurate topographical data.
• Deformation arises from sample instability, substrate adsorption, and tip pressure.

Purpose of the Study:

• To identify experimental conditions that minimize biomolecular deformation during AFM imaging.
• To determine the key factors influencing the accurate height measurement of tobacco mosaic virus (TMV).
• To provide guidelines for optimal AFM imaging of soft biomolecules.

Main Methods:

• Measured the maximum height of tobacco mosaic virus (TMV) under eight different AFM experimental conditions.
• Varied imaging environments (air vs. liquid), substrates (Mica vs. self-assembled monolayers [SAMs]), and imaging modes (tapping mode [TAP] vs. PeakForce tapping [PFT]).
• Used TMV's maximum height as an indicator of sample deformation.

Main Results:

• Substrate type critically affects TMV height measurements in air; SAMs yield more accurate results than Mica.
• Imaging mode is the most critical parameter in liquid environments, with PFT outperforming TAP.
• Optimal TMV height measurements were achieved using PFT in both air and liquid, specifically on SAM substrates.

Conclusions:

• This study clarifies why previous AFM imaging of biomolecules on Mica in air yielded poor results.
• Self-assembled monolayers (SAMs) provide a superior substrate for minimizing biomolecular deformation during AFM.
• PeakForce tapping (PFT) is the recommended imaging mode for accurate AFM topographical analysis of soft biomolecules.