Interlaboratory round robin on cantilever calibration for AFM force spectroscopy

Area of Science:

• Atomic Force Microscopy (AFM)
• Single-Molecule Force Spectroscopy
• Biophysics

Background:

• Accurate cantilever spring constant determination is vital for reliable AFM force spectroscopy.
• Existing calibration methods lack consensus for soft and V-shaped cantilevers.

Purpose of the Study:

• Establish a standardized, accurate protocol for calibrating compliant and V-shaped AFM cantilevers.
• Compare the efficacy of thermal noise and Sader methods for cantilever calibration.

Main Methods:

• Conducted a round-robin experiment across eight laboratories.
• Compared thermal noise and Sader calibration methods on ten AFM systems.
• Utilized both commercial and custom-built AFM cantilevers, including V-shaped designs.

Main Results:

• Both thermal noise and Sader methods accurately determine spring constants for various cantilevers.
• The Sader method demonstrated superior performance for cantilever calibration.
• Simultaneous application of both methods serves as a reliable instrument consistency check.

Conclusions:

• The proposed protocol ensures reliable spring constant determination for diverse AFM cantilevers.
• Accurate calibration is essential, as errors significantly impact biophysical parameter derivation in biological force spectroscopy.
• The Sader method and combined approach offer optimal, reproducible AFM cantilever calibration.