BACKGROUND: Sentinel lymph node (SLN) biopsy is a selective approach to axillary staging of breast cancer with reduced morbidity. Current detection methods including radioisotope and blue dye show good results but some drawbacks are remaining. Indocyanine green (ICG) fluorescence detection was evaluated as a new method for SLN biopsy in breast cancer allowing both transcutaneous visualization of lymphatic vessels and intraoperative identification of SLN. METHODS: Forty-three women with clinically node negative breast cancer received subareolar injection of ICG for fluorescence detection of SLN. All patients underwent either planned axillary lymph node dissection (ALND) with SLN biopsy or selective SLN biopsy to determine need for ALND. Clinical feasibility, detection rate, sensitivity, and axillary recurrence after isolated SLN biopsy were analyzed. RESULTS: Overall ICG fluorescence imaging identified 2.0 SLN in average in 42 of 43 patients (detection rate: 97.7%). Metastatic involvement of the SLN was found in 17 of 18 nodal positive patients by conventional histopathology (sensitivity: 94.4%). Immunohistochemistry revealed isolated tumor cells in five further cases. There was only one false-negative case in 43 patients (5.6%). In 17 of 23 overall nodal positive patients, the SLN was the only positive lymph node. After a median follow-up of 4.7 years none of the patients presented with axillary recurrence. CONCLUSION: ICG fluorescence imaging is a new method for SLN biopsy in breast cancer with acceptable sensitivity and specificity comparable to conventional methods. One advantage of this technique is that it allows transcutaneous visualization of lymphatic vessels and intraoperative lymph node detection without radioisotope.