This review provides a comprehensive overview of the synthesis, biological interactions, and biomedical applications of iron oxide nanoparticles (IONPs). IONPs, known for their magnetic behavior and semiconductor properties, have gained significant attention in various biomedical fields due to their biocompatibility, biodegradability, and low toxicity. However, their clinical use is limited due to insufficient understanding of their biomedical applications. The review highlights the importance of summarizing the preparation techniques, biological interactions in different animal models and cell types, and clinical applications to enhance the safety and effectiveness of IONPs. The synthesis methods for IONPs include chemical, physical, and biological approaches, with chemical methods being the most prevalent. Co-precipitation, micro-emulsion, sol-gel, and thermal decomposition are common chemical methods, while powder ball milling, electron beam lithography, aerosol, and gas phase deposition are physical methods. Biological synthesis methods, such as microbial enzymes and plant phytochemicals, are also discussed. In animal models, the biocompatibility, bio-distribution, metabolism, and bio-clearance of IONPs have been extensively studied. The shape, size, and surface properties of IONPs significantly influence their biological interactions. For example, short rod-shaped IONPs accumulate in the liver, while long rod-shaped IONPs accumulate in the spleen. Size and surface modifications, such as coating with hydrophilic polymers, can improve biocompatibility and reduce toxicity. In vitro studies demonstrate that IONPs can target various types of tumor cells and induce cell death without affecting normal cells. The cytotoxicity of IONPs is influenced by factors such as shape, surface modification, size, concentration, and valence state. External magnetic fields, radiofrequency generator irradiation, MRI imaging, and photothermal therapy can synergistically enhance the anticancer effects of IONPs. IONPs also show promise in non-tumor cells, including osteoblasts, immune cells, and stem cells. They can promote osteoblast differentiation, influence dendritic cell migration, and support stem cell growth and neurite outgrowth. Overall, the review emphasizes the need for further research to optimize the design and application of IONPs in biomedical research and clinical trials, focusing on understanding their biological mechanisms and improving their safety and efficacy.This review provides a comprehensive overview of the synthesis, biological interactions, and biomedical applications of iron oxide nanoparticles (IONPs). IONPs, known for their magnetic behavior and semiconductor properties, have gained significant attention in various biomedical fields due to their biocompatibility, biodegradability, and low toxicity. However, their clinical use is limited due to insufficient understanding of their biomedical applications. The review highlights the importance of summarizing the preparation techniques, biological interactions in different animal models and cell types, and clinical applications to enhance the safety and effectiveness of IONPs. The synthesis methods for IONPs include chemical, physical, and biological approaches, with chemical methods being the most prevalent. Co-precipitation, micro-emulsion, sol-gel, and thermal decomposition are common chemical methods, while powder ball milling, electron beam lithography, aerosol, and gas phase deposition are physical methods. Biological synthesis methods, such as microbial enzymes and plant phytochemicals, are also discussed. In animal models, the biocompatibility, bio-distribution, metabolism, and bio-clearance of IONPs have been extensively studied. The shape, size, and surface properties of IONPs significantly influence their biological interactions. For example, short rod-shaped IONPs accumulate in the liver, while long rod-shaped IONPs accumulate in the spleen. Size and surface modifications, such as coating with hydrophilic polymers, can improve biocompatibility and reduce toxicity. In vitro studies demonstrate that IONPs can target various types of tumor cells and induce cell death without affecting normal cells. The cytotoxicity of IONPs is influenced by factors such as shape, surface modification, size, concentration, and valence state. External magnetic fields, radiofrequency generator irradiation, MRI imaging, and photothermal therapy can synergistically enhance the anticancer effects of IONPs. IONPs also show promise in non-tumor cells, including osteoblasts, immune cells, and stem cells. They can promote osteoblast differentiation, influence dendritic cell migration, and support stem cell growth and neurite outgrowth. Overall, the review emphasizes the need for further research to optimize the design and application of IONPs in biomedical research and clinical trials, focusing on understanding their biological mechanisms and improving their safety and efficacy.