Twist-encoded magnetic plasmon interferometry for label-free, orientation-resolved single-molecule imaging.

Molecular twist-related rotational dynamics at solid-liquid interfaces governs recognition, dissipation, and barrier crossing, yet direct label-free visualization of single-molecule torsion remains challenging. Here, we present twist-encoded magnetic plasmon interferometry (TEMPi), which converts nanorod twist near a metal film into a phase-encoded, orientation-resolved optical signal. Near-field coupling between a gold nanorod and its mirror image in the film excites a localized magnetic plasmon, enabling quantitative mapping of probe twist to interferometric phase. Wide-field, label-free operation in native buffers enables real-time tracking of rotational dynamics and interfacial conformational changes. TEMPi resolves three confinement-dependent rotational regimes, distinguishes effective dual- and single-contact protein anchoring, and localizes single binding sites on the probe through twist-dependent centroid shifts. Applied to DNA hybridization, it quantifies an effective ninefold increase in torsional stiffness and an order-of-magnitude faster twisting relaxation upon duplex formation. These results establish TEMPi as a general twist-resolved metrology for probing effective interfacial molecular mechanics and designing torsion-responsive interfaces in chemistry, biology, and nanotechnology.