In recent decades, targeted drug delivery systems for breast cancer treatment emerged as an ideal alternative and promising solution to reduce systemic side effects of chemotherapeutic agents. In this study, the preparation and characterization of cationic doxorubicin (DOX) loaded magnetic dextran-spermine (DEX-SP) nanocarriers (DEX-SP-DOX) by ionic gelation were fully investigated. Then, anti-HER2 as a monoclonal antibody (mAb) and targeting ligand was conjugated via EDC/NHS reagents. The binding was confirmed by Bradford assay and further assessments were carried out by size and zeta potential measurements. Cytotoxicity effect and internalization of magnetic nanocarriers were assessed by MTT and Prussian blue assays and transmission electron microscopy (TEM), respectively. DLS measurements indicated that the size of nanocarriers increased from 62 to 84 nm by conjugation of anti-HER2 to them. The in vitro release of DOX from mAb conjugated magnetic nanocarriers at pHs 5 and 7.4 was found to be 85 and 55.5%, respectively. The MTT and Prussian blue assays demonstrated enhanced and selective uptake of DEX-SP-DOX-mAb by SKBR cell (HER2 overexpressed cells) in comparison with unconjugated nanocarriers due to higher cellular binding. The TEM result also confirmed cellular internalization of DEX-SP-DOX-mAb magnetic nanocarriers. These results are very promising for targeted delivery of DOX to HER2 positive breast cancer cells.