Magnetic molecular nanorulers enable combinatorial biosensing and determine distances at the molecular level

Molecular nanorulers are essential tools for mapping intramolecular distances and studying nucleic acid and protein interactions. Here, we present MARs (MAgnetic molecular nanoRulers), a fundamentally new class of nanorulers based on measuring the magnetization dynamics of rationally engineered and ultra-responsive colloidal magnetic nanoparticles with magnetic particle spectrometry (MPS). By tuning the grafting density of DNA receptors on MARs, we reveal surface molecular crowding as a simple, yet programmable parameter to tune the response of MARs over a broad range of target DNA concentrations. We show that MARs can distinguish target binding positions along receptor strands by regulating how much the binding effect is translated into the response. Combining their switchable binding-to-response and DNA hairpins, we show that MARs can resolve the full combinatorial space of four short DNA targets (24-1) in a one-design-for-all fashion, with blind assays achieving 80% best-prediction accuracy. By modelling MPS response of MARs using the Fokker-Planck equation, we demonstrate that their response is well predictable. Using biotinylated DNA constructs and calibrating their lengths for different numbers of base pair, we reveal that MARs can determine the distance between bound monostreptavidin and their surface with a dynamic distance range of up to 13 nm and a resolution of 1.3 {+/-} 0.2 nm. Our study establishes a new class of molecular nanorulers for resolving conformational changes in nucleic acids and measuring distances at the molecular level in free solution via an adaptable, simple, and rapid workflow, making MARs highly promising for high throughput screening assays.