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Magnetic nanoparticles (MNPs) with proper surface functionalization have beenextensively applied as labels for magnetic immunoassays, carriers forcontrolled drug/gene delivery, tracers and contrasts for magnetic imaging, etc.Here, we introduce a new biosensing scheme based on magnetic particlespectroscopy (MPS) and the self-assembly of MNPs to quantitatively detect H1N1nucleoprotein molecules. MPS monitors the harmonics of oscillating MNPs as ametric for the freedom of rotational motion, thus indicating the bound statesof MNPs. These harmonics can be readily collected from nanogram quantities ofiron oxide nanoparticles within 10 s. H1N1 nucleoprotein molecule hostsmultiple different epitopes that forms binding sites for many IgG polyclonalantibodies. Anchoring IgG polyclonal antibodies onto MNPs triggers thecross-linking between MNPs and H1N1 nucleoprotein molecules, thereby formingMNP self-assemblies. Using MPS and the self-assembly of MNPs, we achieved thesensitivity of 44 nM (442 pmole) for detecting H1N1 nucleoprotein. In addition,the morphologies and the hydrodynamic sizes of the MNP self-assemblies arecharacterized to verify the MPS results. Different MNP self-assembly modelssuch as classical cluster, open ring tetramer, chain model as well as multimers(from dimer to pentamer) are proposed in this paper. Herein, we claim thefeasibility of using MPS and the self-assembly of MNPs as a new biosensingscheme for detecting ultralow concentrations of target biomolecules, which canbe employed as rapid, sensitive, and wash-free magnetic immunoassays.

Detection of Influenza A Virus Nucleoprotein Through the Self-Assembly  of Nanoparticles in Magnetic Particle Spectroscopy-Based Bioassays: A Method  for Rapid, Sensitive, and Wash-free Magnetic Immunoassays