Magnetically Detected Protein Binding Using Spin-Labeled Slow Off-Rate Modified Aptamers

Recentdevelopments in aptamer chemistry open up opportunitiesfor new tools for protein biosensing. In this work, we present anapproach to use immobilized slow off-rate modified aptamers (SOMAmers)site-specifically labeled with a nitroxide radical via azide-alkyneclick chemistry as a means for detecting protein binding. Proteinbinding induces a change in rotational mobility of the spin label,which is detected via solution-state electron paramagnetic resonance(EPR) spectroscopy. We demonstrate the workflow and test the protocolusing the SOMAmer SL5 and its protein target, platelet-derived growthfactor B (PDGF-BB). In a complete site scan of the nitroxide overthe SOMAmer, we determine the rotational mobility of the spin labelin the absence and presence of target protein. Several sites withsufficiently tight affinity and large rotational mobility change uponprotein binding are identified. We then model a system where the spin-labeledSOMAmer assay is combined with fluorescence detection via diamondnitrogen-vacancy (NV) center relaxometry. The NV center spin-latticerelaxation time is modulated by the rotational mobility of a proximalspin label and thus responsive to SOMAmer-protein binding.The spin label-mediated assay provides a general approach for transducingprotein binding events into magnetically detectable signals.