Chemical functionalization and bioconjugation strategies for atomic force microscope cantilevers

Area of Science:

• Biophysics
• Surface Chemistry
• Microscopy

Background:

• Scanning probe microscopy (SPM) techniques, including atomic force microscopy (AFM), are vital for biophysical research.
• SPM enables measurement of forces at the piconewton level, facilitating single-molecule event observation.
• Secure biomolecule attachment to AFM probes is crucial for accurate force measurements, often requiring covalent linkages.

Purpose of the Study:

• To present strategies and protocols for surface modification of AFM probes for biomolecular attachment.
• To enable robust and reliable single-molecule force spectroscopy experiments.
• To optimize the isolation of biomolecular interactions from surface and tip interference.

Main Methods:

• Surface activation of SPM probe tips (silica, silicon nitride) using gas-phase organosilane deposition.
• Implementation of passivation schemes to prevent non-specific interactions.
• Detailed protocols for various biomolecular immobilization techniques onto functionalized surfaces.

Main Results:

• Established methods for creating strong covalent linkages between biomolecules and AFM probe tips.
• Demonstrated effective strategies for surface modification of hydroxylated surfaces.
• Provided protocols that facilitate the isolation of biomolecular forces during AFM analysis.

Conclusions:

• The presented surface modification strategies are effective for biomolecular attachment to AFM probes.
• These methods enhance the reliability and scope of single-molecule force measurements in biophysics.
• The protocols offer a valuable resource for researchers utilizing AFM in molecular interaction studies.