Magnetic nanoparticles (MNPs) have great potential in various areas such as medicine, cancer therapy and diagnostics, biosensing, and material science. In particular, magnetite (Fe3O4) nanoparticles are extensively used for numerous bioapplications due to their biocompatibility, high saturation magnetization, chemical stability, large surface area, and easy functionalization. This paper describes magnetic nanoparticle physical and biological properties, emphasizing synthesis approaches, toxicity, and various biomedical applications, focusing on the most recent advancements in the areas of therapy, diagnostics, theranostics, magnetic separation, and biosensing.