Surface Functionalization, Nucleic Acid Tether Characterization, and Force Calibration for a Magnetic Tweezers Assay

Magnetic tweezers are a force spectroscopy single-molecule technique. They enable the mechanical manipulation of biomolecules via the means of a magnetic particle under an attractive force applied by a magnetic field source. The magnetic particle is tethered to the glass surface of a flow chamber by the biomolecule, and functionalization strategies have been developed to reduce the nonspecific interactions of either the magnetic particles or biomolecules with the surface. Here, we describe two complementary strategies to achieve a high tether density while reducing the interactions of both the magnetic particle and the biomolecule of interest with the glass surface. Using a large detector CMOS camera, the simultaneous observation of several hundreds of tethered magnetic beads is achievable, allowing high-throughput single-molecule measurements. We further describe here a simple procedure to perform the calibration in force of a magnetic tweezers assay.