Using chitosan-modified magnetite nanoparticles (CS-MNPs) as a replacement of enzymes in conventional ELISA configurations, a magnetic nanoparticle-linked immunosorbent assay is demonstrated. The CS-MNPs are synthesized by using a one-step solvothermal process, where chitosan in the reaction system is used both as a ligand and as a surface modification agent. The as-synthesized CS-MNPs have amine groups on their surface which provide good dispersibility in aqueous solutions and convenient sites for covalent linking of antibodies with the MNPs. They also possess catalytic properties that are able to catalyze color reactions in immunoassays, and magnetic properties with both a large enough saturation magnetization and a superparamagneticity that can be used to capture, separate, and enrich antigens prior to the assay procedure. By employing both the catalytic and magnetic properties of the CS-MNPs, a capture-detection immunoassay is developed, where antigens can be captured, separated, and enriched prior to the assay procedure. It is shown that such a procedure enabled by the intrinsic dual functionalities of MNPs may improve the detection limit of immunoassays.