Magnetite nanoparticles (MNPs) grafted with poly(2-acrylamido-2-methylpropanesulfonic acid) (PAMPS) and their use as anionic nano-adsorbents for positively charged antibody were herein presented. PAMPS grafted on MNP surface provided good water dispersibility and high negatively charged surface for electrostatic adsorption with positively charged entities, and MNPs provided a good magnetic responsiveness to facilitate the separation process. From transmission electron microscopy results, the size of the nanoclusters ranged between 50 and 200 nm in diameter with ca. 50–150 particles/cluster. Degree of the negative charge of PAMPS-grafted MNPs increased when increasing pH of the dispersion, indicating the pH-responsive properties of the MNPs. These MNPs were successfully used as nano-adsorbents for adsorption with anti-rabbit IgG through ionic interaction between negative charge of PAMPS-grafted MNP and positive charge of the antibody. In addition, the MNPs also provided the reusable ability by adsorption-desorption cycling process with higher than 80% adsorption ability after five consecutive reusing cycles. These hybrid materials should be alternative nano-adsorbents for other positively charged entities with high adsorption efficiency and reusable supports in magnetic separation applications.