Calcium　doped　magnetite　nanoparticles　were　synthesized　by　the　chemical　co-precipitation　method　for　osteogenetic　and　antibacterial　applications　in　biomedical　fields.　The　prepared　powders　are　composed　of　Fe3O4　and　calcium　ferrite,　but　Fe2O3·H2O　(i.e.　FeOOH)　and　CaCO3　are　also　detected　at　high　calcium　content.　Aggregated　spherical　nanoparticles　(diameter　12–15　nm)　are　observed　by　SEM　and　TEM,　but　clusters　of　needle-like　nanorods　appear　especially　for　samples　with　high　calcium　content.　The　particles　have　much　larger　hydrodynamic　diameter　values　(170–250　nm)　and　positive　surface　charge.　Magnetic　properties　of　the　samples　generally　decrease　with　calcium　content,　including　saturation　magnetization,　coercivity　and　residual　magnetization.　Antibacterial　tests　by　plate　counting,　TEM　observation　and　intracellular　reactive　oxygen　species　(ROS)　assay　suggest　that　the　samples　with　high　calcium　content　have　better　bactericidal　activity,　and　the　main　mechanisms　include　the　physical　mutilation　effect　of　nanoparticles　and　the　oxidative　stress　induced　in　the　bacteria.　Cell　tests　using　MC3T3-E1　cells　show　that　the　tested　samples　have　good　cytocompatibility,　and　the　alkaline　phosphatase　(ALP)　activity　is　dose-dependent　for　the　nanoparticles.　The　developed　Ca-doped　magnetite　nanoparticles　with　high　magnetism,　good　antibacterial　and　osteogenetic　properties　would　find　potential　application　in　different　biomedical　fields.　©　2022　Elsevier　Ltd　and　Techna　Group　S.r.l.