BACKGROUND The sentinel lymph node (SLN) biopsy technique using superparamagnetic iron oxide (SPIO) as a tracer instead of radioisotopes has been described. To further advance this technique, we evaluated preoperative SPIO-MR sentinel lymphography to facilitate the accurate identification of the lymphatic pathways and primary SLN. METHODS A prospective study was performed in ten patients with breast cancer and clinically negative axillary lymph nodes. None of the patients received preoperative chemotherapy. After 1.6 ml of SPIO (ferucarbotran) was injected in the subareolar breast tissue, sentinel axillary lymph nodes were detected by MRI in T2*-weighted gradient echo images and resected using the serial SPIO-SLN biopsy procedure with a handheld magnetometer. RESULTS In one patient, gadolinium-enhanced MR imaging was performed at the same time as SPIO-MR lymphography, and this patient was excluded from further analysis. In all patients (9/9) SLNs were detected by SPIO-MR sentinel lymphography and successfully identified at surgery. The number of SLNs detected by lymphography (mean 2.7) significantly correlated with SLNs identified at surgery (mean 2.2). One patient had nodal metastases. In one patient, skin color changed to brown at the injection site and resolved spontaneously. There were no severe reactions to the procedure or complications in any patient. CONCLUSIONS This is the first study to evaluate SPIO both as a contrast material in MR sentinel lymphography and as a tracer in SLN biopsy using an integrated method. The acquired three-dimensional imaging demonstrated excellent image quality and usefulness to identify SLN in conjunction with SLN biopsy.