The past decade has seen tremendous progress in the synthesis and surface functionalization of iron oxide nanoparticles (IONPs) for a variety of biomedical applications. However, there is still a growing demand on magnetic nanoparticles with higher magnetic moments for more sensitive diagnosis and lower dose treatments in magnetic bioassays, imaging, and therapies. In view of this need, the γ′-Fe 4 N nanoparticle, with around 3 times higher saturation magnetizations than IONPs, becomes one promising alternative for these applications. However, the large and non-uniformly distributed sizes of γ′-Fe 4 N nanoparticles hinder the biomedical applications. These synthesized γ′-Fe 4 N nanoparticles are not suitable for biomedical applications at the current stage. Herein, we have developed and demonstrated a wet ball milling method along with different surface-active media to produce ultrastable, monodispersed, uniformly sized, and sub-100 nm γ′-Fe 4 N nanoparticles in solvents. Different standard characterization methods such as transmission electron microscopy, nanoparticle tracking analysis, and Fourier-transform infrared spectroscopy are carried out to measure the physicochemical properties of these surface-functionalized γ′-Fe 4 N nanoparticles. It is confirmed that the functional chemical groups have been successfully anchored on our purified sub-100 nm γ′-Fe 4 N nanoparticles, which allows for convenient subsequent conjugation of proteins, nucleic acids, and drugs for future in vitro and in vivo biomedical applications.